Pre-clinical development of a hydrogen peroxide-inactivated West Nile virus vaccine.

West Nile virus (WNV) is a mosquito-transmitted pathogen with a wide geographical range that can lead to long-term disability and death in some cases. Despite the public health risk posed by WNV, including an estimated 3 million infections in the United States alone, no vaccine is available for use in humans. Here, we present a scaled manufacturing approach for production of a hydrogen peroxide-inactivated whole virion WNV vaccine, termed HydroVax-001WNV. Vaccination resulted in robust virus-specific neutralizing antibody responses and protection against WNV-associated mortality in mice or viremia in rhesus macaques (RM). A GLP-compliant toxicology study performed in rats demonstrated an excellent safety profile with clinical findings limited to minor and transient irritation at the injection site. An in vitro relative potency (IVRP) assay was developed and shown to correlate with in vivo responses following forced degradation studies. Long-term in vivo potency comparisons between the intended storage condition (2-8°C) and a thermally stressed condition (40±2°C) demonstrated no loss in vaccine efficacy or protective immunity over a 6-month span of time. Together, the positive pre-clinical findings regarding immunogenicity, safety, and stability indicate that HydroVax-001WNV is a promising vaccine candidate.

Glenoid avulsion of the glenohumeral ligament (GAGL): a case report and review of the anatomy.

Shoulder dislocations are frequently seen in the general population and can be a cause of instability. Instability can lead to debilitating symptoms and morbidity as a result of progressive damage to the shoulder. Anterior shoulder dislocations are the most frequent type of dislocations and have been studied extensively with MRI. The soft tissue Bankart lesion is the most well-known entity associated with anterior instability; however, additional structural lesions arising from traumatic events have been described in recent literature which also predispose to anterior shoulder instability. One of these lesions, the glenoid avulsion of the glenohumeral ligament (GAGL), involves avulsion of the inferior glenohumeral ligament from the glenoid and involves separation from an intact labrum. In contrast to the Bankart lesion, there has been limited discussion of the GAGL lesion in the literature and very few imaging examples. We report a case of a GAGL diagnosed on MRI and confirmed with arthroscopy. It is discussed in the context of the anatomy of the inferior glenohumeral ligament and the imaging findings.

The prevalence of chondrocalcinosis of the symphysis pubis on CT scan and correlation with calcium pyrophosphate dihydrate crystal deposition disease.

Calcium pyrophosphate dihydrate (CPP) crystal deposition in the articular cartilage can often be seen radiographically as chondrocalcinosis (CC). CPP crystals preferentially deposit in fibrocartilages such as the knee menisci and symphysis pubis (SP). We sought to determine the prevalence of CC in the SP on computed tomography (CT) of the abdomen and pelvis. This retrospective study involved readings on 1070 consecutive CTs of the abdomen and pelvis performed over 3 months in patients over 65 years of age. Medical records of 226 patients found to have CC were reviewed to determine age, gender, documentation of CPPD on problem lists or in medical histories, and whether radiology readings of the CTs mentioned CC. SP CC was identified in 21.1 % (226/1070) of consecutive CT scans with the mean age of CT+ patients being 78.6. Of the 226 patients with SP CC, the observation of CC was documented in only 5.3 % (12/226) of the radiology reports. Of the 12 instances in which the radiology reports mentioned CC, this observation was never (0/12) transmitted to the medical history or problem list. The prevalence of SP CC in patients older than 65 was 21.1 %. Since the majority of CTs of the abdomen and pelvis are not ordered for evaluation of musculoskeletal conditions, this is likely a true prevalence without selection bias. When CC of the SP was present on images, radiologists routinely overlooked or chose not to report CC. Even in the rare instances when it was reported, that information was not added to the medical history or problem list. There are several clinical situations (e.g., acute monoarthritis or atypical osteoarthritis) in which recognizing that a patient has CPP deposition would be useful. Taking the time to review images may yield clinically important findings that are not mentioned anywhere on the patient chart.

Iliotibial band avulsion fracture: a case report with differential diagnosis.

Avulsion injuries of the knee are common sequelae of significant trauma given the number of ligamentous and tendinous insertions around the joint. Commonly discussed avulsion fractures of the lateral knee include the Segond fracture of the lateral tibial plateau and the arcuate complex avulsion fracture of the fibular styloid process. A less common avulsion fracture is the iliotibial (IT) band avulsion fracture involving the anterolateral corner of the tibia (Gerdy's tubercle). It is crucial to identify IT band avulsion fractures because of the frequent associated internal derangements of the knee. This case report describes the imaging of an acute IT band avulsion fracture and compares these findings with other lateral knee avulsion fractures.

Complications in musculoskeletal intervention: important considerations.

Musculoskeletal (MSK) intervention has proliferated in recent years among various subspecialties in medicine. Despite advancements in image guidance and percutaneous technique, the risk of complication has not been fully eliminated. Overall, complications in MSK interventions are rare, with bleeding and infection the most common encountered. Other complications are even rarer. This article reviews various complications unique to musculoskeletal interventions, assists the reader in understanding where pitfalls lie, and highlights ways to avoid them.

Practice policy and quality initiatives: using lean principles to improve screening mammography workflow.

The "lean" approach is a quality improvement method that focuses on maximizing activities that are valued by the customer and eliminating waste that impedes efficiency in the workplace. The unique philosophy of the lean approach encourages all members of the team to be directly involved in identifying areas of waste and generating solutions to eliminate them. When the breast imaging section at the authors' institution became part of a multispecialty breast care center, the result was escalating examination volumes, more complex cases, and overall increased demand on radiologists' time. After several unsuccessful attempts to improve the efficiency of the section, including evaluation by outside consultants, the decision was made to embark on a comprehensive quality improvement program using the lean approach. A team of radiologists, technologists, file room personnel, information technology (IT) representatives, and administrators from the breast imaging section met twice a month to learn about lean principles and how to apply them to screening mammography workflows. Sources of inefficiency (waste) were identified, and potential solutions were generated. Multiple trials were performed to test these solutions. Throughout the process, all team members were engaged in identifying the problems, suggesting solutions, and implementing change. Most of the tested solutions were successful and resulted in decreased patient wait times, improved efficiency for the technologists and radiologists, faster report turnaround, and advances in IT. In addition, staff members were introduced to the lean philosophy and became actively involved in improving their workplace, resulting in a more cohesive section.

The prevalence of chondrocalcinosis (CC) of the acromioclavicular (AC) joint on chest radiographs and correlation with calcium pyrophosphate dihydrate (CPPD) crystal deposition disease.

Digital imaging combined with picture archiving and communication system (PACS) access allows detailed image retrieval and magnification. Calcium pyrophosphate dihydrate (CPPD) crystals preferentially deposit in fibrocartilages, the cartilage of the acromioclavicular (AC) joint being one such structure. We sought to determine if examination of the AC joints on magnified PACS imaging of chest films would be useful in identifying chondrocalcinosis (CC). Retrospective radiographic readings and chart reviews involving 1,920 patients aged 50 or more who had routine outpatient chest radiographs over a 4-month period were performed. Knee radiographs were available for comparison in 489 patients. Medical records were reviewed to abstract demographics, chest film reports, and diagnoses. AC joint CC was identified in 1.1 % (21/1,920) of consecutive chest films. Patients with AC joint CC were 75 years of age versus 65.4 in those without CC (p < 0.0002). Four hundred eighty-nine patients had knee films. Six of these patients had AC joint CC, and of these, five also had knee CC (83 %). Of the 483 without AC joint CC, 62 (12 %) had knee CC (p = 0.002). Patients with AC joint CC were more likely to have a recorded history of CPPD crystal deposition disease than those without AC joint CC (14 versus 1 %, p = 0.0017). The prevalence of AC joint CC increases with age and is associated with knee CC. A finding of AC joint CC should heighten suspicion of pseudogout or secondary osteoarthritis in appropriate clinical settings and, in a young patient, should alert the clinician to the possibility of an associated metabolic condition.

A flow cytometry-based assay for quantifying non-plaque forming strains of yellow fever virus.

Primary clinical isolates of yellow fever virus can be difficult to quantitate by standard in vitro methods because they may not form discernable plaques or induce a measurable cytopathic effect (CPE) on cell monolayers. In our hands, the Dakar strain of yellow fever virus (YFV-Dakar) could not be measured by plaque assay (PA), focus-forming assay (FFA), or by measurement of CPE. For these reasons, we developed a YFV-specific monoclonal antibody (3A8.B6) and used it to optimize a highly sensitive flow cytometry-based tissue culture limiting dilution assay (TC-LDA) to measure levels of infectious virus. The TC-LDA was performed by incubating serial dilutions of virus in replicate wells of C6/36 cells and stained intracellularly for virus with MAb 3A8.B6. Using this approach, we could reproducibly quantitate YFV-Dakar in tissue culture supernatants as well as from the serum of viremic rhesus macaques experimentally infected with YFV-Dakar. Moreover, the TC-LDA approach was >10-fold more sensitive than standard plaque assay for quantitating typical plaque-forming strains of YFV including YFV-17D and YFV-FNV (French neurotropic vaccine). Together, these results indicate that the TC-LDA technique is effective for quantitating both plaque-forming and non-plaque-forming strains of yellow fever virus, and this methodology may be readily adapted for the study and quantitation of other non-plaque-forming viruses.

Optimization of peptide-based ELISA for serological diagnostics: a retrospective study of human monkeypox infection.

Although smallpox has been eradicated, other diseases caused by virulent orthopoxviruses such as monkeypox virus (MPV) remain endemic in remote areas of western and central sub-Saharan Africa, and represent a potential biothreat due to international travel and/or inadvertent exposure. Unfortunately, extensive antigenic cross-reactivity among orthopoxviruses presents a challenge to serological diagnosis. We previously reported a 20mer peptide-based ELISA that identified recent MPV infection with >90% sensitivity and >90% specificity. However, the sensitivity of this approach was not determined with samples obtained at later time points after antibody titers had declined from their peak levels. To improve assay sensitivity for detecting MPV-specific antibodies at later time points, we compared diagnostic 20mer peptides to 30mer peptides. In addition, optimal 30mer peptides were tested in combination or after conjugating selected peptides to a carrier protein (bovine serum albumin) to further improve assay performance. An optimized combination of four unconjugated 30mer peptides provided 100% sensitivity for detecting MPV infection at 2-6 months post-infection, 45% sensitivity for detecting MPV infection at >2 years post-infection, and 99% specificity. However, an optimized combination of two peptide conjugates provided 100% sensitivity for detecting MPV infection at 2-6 months post-infection, 90% sensitivity for detecting MPV infection at >2 years post-infection, and 97% specificity. Peptide-based ELISA tests provide a relatively simple approach for serological detection of MPV infection. Moreover, the systematic approach used here to optimize diagnostic peptide reagents is applicable to developing improved diagnostics to a broad range of other viruses, and may be particularly useful for distinguishing between closely-related viruses within the same genus or family.

Osteoporosis screening and treatment guidelines: are they being followed?

OBJECTIVE: The objective of this study was to examine a cohort of women sent for dual-energy x-ray absorptiometry (DXA) screening to see whether they met the criteria for bone density testing. In addition, we sought to determine whether they were receiving appropriate interventions, based on published guidelines. METHODS: Between January 1, 2007, and March 1, 2009, inclusive, postmenopausal women (age >49 y) who were sent for DXA bone density screening were offered enrollment into the study. Risk factors for osteoporosis, demographic information, and current DXA results were recorded. The 2006 Osteoporosis Position Statement of The North American Menopause Society was used for screening and therapeutic intervention guidelines. RESULTS: Among the 615 women with data, the mean (SD) age was 61.5 (8.3) years. Using the 2006 guidelines of The North American Menopause Society, 41.3% (253 of 612) of the women who had DXA testing did not meet the criteria for such screening. Of these women, 25.5% (157 of 615) were not taking calcium, 31.1% (191 of 614) were not taking vitamin D, and 59.8% (343 of 574) were not exercising at least half an hour per week. Of the women with any of the approved indications for treatment, 15.7% (16 of 102) were not taking calcium, 18.6% (19 of 102) were not taking vitamin D, 52.7% (49 of 93) were not exercising at least 2 hours per week, and 35.3% (36 of 102) were not receiving therapy. In contrast, of those women without an indication for treatment, 17.8% (83 of 467) were receiving bisphosphonate, raloxifene, or calcitonin therapy. CONCLUSIONS: A large number of women are not properly screened or treated for osteoporosis. Inappropriate screening may also lead to improper management of osteoporosis and its associated complications.

Fabella fracture with CT imaging: a case report.

Fracture of the fabella is rare, may be easily overlooked, and can be a clinically important cause of posterolateral knee pain following traumatic injury or total knee arthroplasty. To date, nine case reports of fabella fracture with radiographic documentation have been reported in the literature. This report documents a 55-year-old male pedestrian who was struck by an automobile and presented with radiographs demonstrating depressed lateral tibial plateau and proximal fibula fractures. Computed tomography (CT) was performed for surgical planning and demonstrated the additional finding of a radiographically occult nondisplaced fabella fracture. To the best of our knowledge, this is the first case in which CT documentation of a fabella fracture is reported. Fracture of the fabella is a rare but important clinical entity which may be overlooked clinically and radiographically. Clinical information can provide a high index of suspicion, and when coupled with radiographic and CT findings, may lead to the correct diagnosis. CT imaging of the knee may confirm a suspected fabella fracture or may help detect a radiographically occult fracture.

Serologic evidence for novel poxvirus in endangered red Colobus monkeys, Western Uganda.

Enzyme-linked immunosorbent assay, Western blot, and virus neutralization assays indicated that red colobus monkeys in Kibale National Park, western Uganda, had antibodies to a virus that was similar, but not identical, to known orthopoxviruses. The presence of a novel poxvirus in this endangered primate raises public health and conservation concerns.

Retrospective analysis of monkeypox infection.

Serologic cross-reactivity between orthopoxviruses is a substantial barrier to laboratory diagnosis of specific orthopoxvirus infections and epidemiologic characterization of disease outbreaks. Historically, time-consuming and labor-intensive strategies such as cross-adsorbed neutralization assays, immunofluorescence assays, and hemagglutination-inhibition assays have been used to identify orthopoxvirus infections. We used cross-adsorption to develop a simple and quantitative postadsorption ELISA for distinguishing between monkeypox and vaccinia infections. Despite the difficulty of diagnosing clinically inapparent monkeypox in previously vaccinated persons, this technique exhibited 100% sensitivity and 100% specificity for identifying clinically overt monkeypox infection irrespective of vaccination history. We also describe a Western blot technique in which up to 3 diagnostic bands may be used to distinguish between vaccinia and monkeypox infection. The techniques described provide independent diagnostic tests suitable for retrospective analysis of monkeypox outbreaks.

Trypanosomes expressing a mosaic variant surface glycoprotein coat escape early detection by the immune system.

Host resistance to African trypanosomiasis is partially dependent on an early and strong T-independent B-cell response against the variant surface glycoprotein (VSG) coat expressed by trypanosomes. The repetitive array of surface epitopes displayed by a monotypic surface coat, in which identical VSG molecules are closely packed together in a uniform architectural display, cross-links cognate B-cell receptors and initiates T-independent B-cell activation events. However, this repetitive array of identical VSG epitopes is altered during the process of antigenic variation, when former and nascent VSG proteins are transiently expressed together in a mosaic surface coat. Thus, T-independent B-cell recognition of the trypanosome surface coat may be disrupted by the introduction of heterologous VSG molecules into the coat structure. To address this hypothesis, we transformed Trypanosoma brucei rhodesiense LouTat 1 with the 117 VSG gene from Trypanosoma brucei brucei MiTat 1.4 in order to produce VSG double expressers; coexpression of the exogenous 117 gene along with the endogenous LouTat 1 VSG gene resulted in the display of a mosaic VSG coat. Results presented here demonstrate that the host's ability to produce VSG-specific antibodies and activate B cells during early infection with VSG double expressers is compromised relative to that during infection with the parental strain, which displays a monotypic coat. These findings suggest a previously unrecognized mechanism of immune response evasion in which coat-switching trypanosomes fail to directly activate B cells until coat VSG homogeneity is achieved. This process affords an immunological advantage to trypanosomes during the process of antigenic variation.

Antibody-mediated protection against respiratory viral infection.

Effective antibody responses are critical for protection against many human pathogens, including those with tropism for the respiratory tract (RT). Systemic immunoglobulin (Ig)G responses as well as mucosal IgA responses play complementary roles in protection against RT infections, and induction of a tissue-specific, isotype-appropriate humoral response is central to successful vaccination strategies. For respiratory virus infections in which current vaccines are either not available or not fully effective, antibody-mediated therapies may provide a viable treatment option. Prophylactic administration of polyclonal or monoclonal antibodies shows the best clinical efficacy, whereas therapeutic administration of antibodies after infection can also be highly protective but is greatly dependent on timing; efficacy declines soon after onset of disease symptoms. Further understanding of the mechanisms underlying antibody-mediated protection against respiratory virus infections may lead to improved immunization strategies as well as more effective antibody-based therapeutics.