Comparison of Rodent Infectious Agent Detection by Exhaust Dust Testing and Traditional Sentinel Testing Using Quantitative Polymerase Chain Reaction.

Improved diagnostic capabilities and a desire to reduce or refine the use of animals as soiled bedding sentinels (SBS) have driven interest in developing the use of PCR-based testing methods, such as exhaust dust testing (EDT), for routine rodent health surveillance. We compared the absolute and quantitative PCR results from EDT filters with SBS mice by routine screening via a panel of 19 infectious agents including agents known to be excluded or present in the colony. In this study, EDT and SBS were compared at days 0, 90, and 180 in 3 facilities (n = 12 rooms) with animals housed on IVC racks (n = 19 double-sided and 23 single-sided racks). All racks were negative for excluded agents (n = 15 agents) during the study. The bacterial agent Helicobacter spp. was consistently detected on EDT filters while less consistently detected via SBS. EDT filters detected Corynebacterium bovis better than SBS in areas where the agent was present. EDT filters and SBS mice tested for murine norovirus (MNV) demonstrated agreement for positive tests by both PCR and serology. For rodent chaphamaparvovirus-1 (RCHPV-1) we compared EDT to urine and feces from SBS. Six cages of SBS were positive for RCHPV-1 by fecal PCR with 5 out of 6 testing positive on urine, while only 3 out of 6 EDT filters tested positive. Since real-time fluorogenic PCR was used for testing, relative PCR copy numbers for each positive finding were evaluated to estimate organism load at the rack level. Copy numbers allowed for further characterization of agent presence within a colony. Furthermore, we compared copy numbers with cage census for MNV and Helicobacter spp., which was positively correlated for EDT testing but not for SBS. Overall, our results demonstrate that EDT's ability to detect many commonly excluded agents is comparable to or better than SBS.

Comparison of Plenum and Cage-level Filter Exhaust Dust PCR Testing to Soiled Bedding Sentinel Mice (Mus musculus) on an IVC Rack.

The use of soiled-bedded sentinels (SBSs) has historically been the standard for colony health surveillance monitoring at our institution. With the advent of newer technologies in which dust collected from filters is tested by PCR, we compared traditional SBS with PCR testing of both exhaust air dust collected from a filter in the downstream vertical plenum (exhaust dust test [EDT]) and the SBS cage-level exhaust filter (SCEF). Our hypothesis was that both methods of filter testing would identify more pathogens than SBS testing. Twenty-five individually ventilated mouse racks that used disposable caging were sanitized and placed into rotation. Rack plenums were tested by PCR to verify negative results before the study start. Exhaust dust collection media were placed in the exhaust plenum (n = 25). SBS cages were placed on each side of the rack with 2 mice per cage (n = 42 mice), with the remaining cage slots occupied by research animals. At each triweekly cage change, the exhaust air filters were carefully removed from the cage top, placed in sterile 50-mL conical tubes, and pooled for submission. After 3mo, the SBS mice were tested via serology for bacterial and viral agents and by PCR for Helicobacter species, pinworms, and ectoparasites. In addition, the EDT filter and SCEF were collected for PCR to evaluate for the same agents. Our results indicate that the SCEF consistently detected agents more frequently than the EDT filter placed in the plenum and that the EDT filter media detected agents more frequently than did the SBS mice. Our data suggest that both PCR methods of detection are superior to SBS for individually ventilated disposable rodent cages and that the SCEF is superior to EDT. These data supported our movement of institution toward environmental monitoring as a method of rodent colony health surveillance.

Evaluation of a Novel Battery-Operated Tumbler Device for Use in the Detection of Mouse Pathogens for Rodent Health Monitoring.

The search for alternatives to live animal sentinels in rodent health monitoring programs is fundamental to the 3Rs (Reduction, Replacement, and Refinement) of animal research. We evaluated the efficacy of a novel battery-operated tumbler device that rotates soiled bedding in direct contact with sample media against the use of exhaust sample media and soiled bedding sentinel (SBS) mice. Four rodent racks were used, each with 3 test cages: a cage with a tumbler device that rotated for 10min twice a week (TUM10), a cage with a tumbler device that rotated for 60min twice a week (TUM60), and a cage housing 2 female Crl:CD1(ICR) mice. Every 2 wk, each test cage received soiled bedding collected from all cages on each respective rack. In addition to soiled bedding, the tumbler device contained various sample collection media: a contact Reemay filter (3mo-cRF) that remained in the tumbler for the duration of the study, a contact Reemay filter (1mo-cRF) that was replaced monthly, adhesive swabs (AS) that were added at every biweekly cage change, and an exhaust Reemay filter located at the exhaust outlet of the cage. All analyses were performed by direct PCR for both sample media in the animal-free methods, and fecal pellet, body swab, and oral swabs were collected from sentinel mice. Out of 16 total pathogens detected, assessment of 1mo-Crf from both TUM10 and TUM60 cages detected 84% and 79% of pathogens, respectively, while SBS samples detected only 47% of pathogens. AS in TUM60 and TUM10 cages detected the fewest pathogens (24% and 13%, respectively). These results indicate that the novel tumbler device is an effective and reliable tool for rodent health monitoring programs and a suitable replacement for live animal sentinels. In this study, 1mo-cRF in TUM10 cages detected the highest number of pathogens.

Using Filter Media and Soiled Bedding in Disposable Individually Ventilated Cages as a Refinement to Specific Pathogen-free Mouse Health Monitoring Programs.

Molecular-based methods have shown potential for improving pathogen detection and reducing animal use. While increasing evidence supports rodent-free environmental health PCR pathogen detection, limited information is available regarding efficacy for disposable individually ventilated caging systems. In such systems, testing of plenum exhaust air dust is ineffective, and the use of collection media is optimal. We performed a series of studies to compare PCR infectious agent detection with dust collected on media placed in a mouse-free soiled bedding cage, the cage exhaust filter of an occupied sentinel cage, and direct sampling from colony and sentinel mice with traditional soiled bedding mouse sentinels. We hypothesized that after a 3-mo period, testing of filter media agitated in a soiled bedding cage would be equal to or more sensitive than more traditional methods. Agitated media detected Astrovirus-1, segmented filamentous bacteria and Helicobacter ganmani to a degree comparable to testing lid exhaust filter PCR from a sentinel mouse cage, but opportunists such as Staphylococcus aureus and Proteus mirabilis were not detected consistently, and H. hepaticus was not detected at all. Direct sampling of pooled fecal pellets and body swabs from sentinel mice and testing using PCR also failed to reliably detect opportunists and Helicobacter spp. While further work is needed to refine use of filter media in soiled bedding for detection of lower prevalence opportunists, this report provides evidence that a rodent-free method of reliably detecting murine agents in a disposable individually ventilated cage system with cage-level filtration outperforms direct sampling of soiled bedding sentinel mice.

Assessing Elimination of Mouse Kidney Parvovirus from Cages by Mechanical Washing.

Mouse kidney parvovirus (MKPV), a newly identified parvovirus of the genus Chaphamaparvovirus, causes inclusion body nephropathy in severely immunocompromised mice and is prevalent in research mouse colonies. As nonenveloped viruses, mammalian parvoviruses are stable and generally resist thermal inactivation; however, as a novel and highly divergent parvovirus, the thermal stability of MKPV is undefined. This study aimed to evaluate the ability of cage sanitization in a mechanical washer to eliminate MKPV. Cages contaminated by MKPV-infected mice were assigned to 1 of 3 treatment groups: 1) control (bedding change only); 2) sanitization in a tunnel washer (88°C final rinse for 20 s); or 3) sanitization in a tunnel washer followed by autoclave sterilization (121 °C for 20 min). The presence of MKPV on the cage's interior surface was assessed by PCR of cage swab extracts collected before and after cage treatment. After treatment and swabbing, each cage housed 4 MKPV-negative CD1 mice. Each group of naive CD1 mice was assigned to one of the treatment groups and was housed in a cage from this group for two, 1 wk periods. At 12, 17, and 20 wk after the first exposure, renal tissue was collected from 1 test mouse per cage and assessed for MKPV by PCR. MKPV was detected by PCR on the surface of 63% of the pretreatment cages. All cages sanitized in a tunnel washer with or without sterilization were PCR negative after treatment. Seven of 10 mice housed in untreated cages contained a mouse positive for MKPV by 20 wk after exposure. None of the mice housed in cages sanitized in a tunnel washer with or without sterilization tested positive for MKPV at any time point. This study indicates that MKPV contaminated caging can result in MKPV infection of mice, and the use of a tunnel washer at the temperature and duration evaluated was sufficient to remove MKPV nucleic acid and prevent MKPV transmission.

The Effects of Water Volume and Bacterial Concentration on the Water Filtration Assay Used in Zebrafish Health Surveillance.

The number of zebrafish in biomedical research has increased exponentially over the past decades, leading to pressure on the laboratory animal community to develop and refine techniques to monitor zebrafish health so that suitable stocks can be maintained for research. The water filtration assay is a promising technique in which water from a zebrafish system is filtered, and the filter analyzed by PCR. In the present report, we studied how the volume of water tested and the concentration of bacterial pathogens affected test results. To do so, we used stock solutions of 3 zebrafish pathogens: Edwardsiella ictaluri, Aeromonas hydrophila, and Mycobacterium marinum. We used these stocks to create solutions with known concentrations of each pathogen, ranging between 10² and 107 Colony Forming Units (CFU) per ml. One, 2, and 3 L of each solution was filtered using positive pressure, and the filters were submitted to a commercial lab for PCR testing. Results were fit with a logistic regression model, and the probability of obtaining a positive result were calculated. Test sensitivity varied by organism, but in general, test results were positively correlated with the volume of the water filtered and with the concentration of bacteria in solution. We conclude that a positive result can be expected for E. ictaluri at 105 CFU per mL, A. hydrophila at 106 CFU per ml, and M. marinum at 106 CFU per mL, when 3 L of solution are filtered.

Chemotherapy and Targeted Therapy for Patients With Human Epidermal Growth Factor Receptor 2-Negative Metastatic Breast Cancer That is Either Endocrine-Pretreated or Hormone Receptor-Negative: ASCO Guideline Update.

PURPOSE: This guideline updates recommendations of the ASCO guideline on chemotherapy and targeted therapy for patients with human epidermal growth factor receptor 2-negative metastatic breast cancer (MBC) that is either endocrine-pretreated or hormone receptor (HR)-negative. METHODS: An Expert Panel conducted a targeted systematic literature review guided by a signals approach to identify new, potentially practice-changing data that might translate into revised guideline recommendations. RESULTS: The Expert Panel reviewed abstracts from the literature review and retained 14 articles. RECOMMENDATIONS: Patients with triple-negative, programmed cell death ligand-1-positive MBC may be offered the addition of immune checkpoint inhibitor to chemotherapy as first-line therapy. Patients with triple-negative, programmed cell death ligand-1-negative MBC should be offered single-agent chemotherapy rather than combination chemotherapy as first-line treatment, although combination regimens may be offered for life-threatening disease. Patients with triple-negative MBC who have received at least two prior therapies for MBC should be offered treatment with sacituzumab govitecan. Patients with triple-negative MBC with germline BRCA mutations previously treated with chemotherapy may be offered a poly (ADP-ribose) polymerase inhibitor rather than chemotherapy. Patients with HR-positive human epidermal growth factor receptor 2-negative MBC for whom chemotherapy is being considered should be offered single-agent chemotherapy rather than combination chemotherapy, although combination regimens may be offered for highly symptomatic or life-threatening disease. Patients with HR-positive MBC with disease progression on an endocrine agent may be offered treatment with either endocrine therapy with or without targeted therapy or single-agent chemotherapy. Patients with HR-positive MBC with germline BRCA mutations no longer benefiting from endocrine therapy may be offered a poly (ADP-ribose) polymerase inhibitor rather than chemotherapy. No recommendation regarding when a patient's care should be transitioned to hospice or best supportive care alone is possible.Additional information is available at www.asco.org/breast-cancer-guidelines.

Selection of Optimal Adjuvant Chemotherapy and Targeted Therapy for Early Breast Cancer: ASCO Guideline Update.

PURPOSE: The aim of this work is to update key recommendations of the ASCO guideline adaptation of the Cancer Care Ontario guideline on the selection of optimal adjuvant chemotherapy regimens for early breast cancer and adjuvant targeted therapy for breast cancer. METHODS: An Expert Panel conducted a targeted systematic literature review guided by a signals approach to identify new, potentially practice-changing data that might translate into revised guideline recommendations. RESULTS: The Expert Panel reviewed abstracts from the literature review and identified one article for inclusion that reported results of the phase III, open-label KATHERINE trial. In the KATHERINE trial, patients with stage I to III human epidermal growth factor receptor 2 (HER2)-positive breast cancer with residual invasive disease in the breast or axilla after completing neoadjuvant chemotherapy and HER2-targeted therapy were allocated to adjuvant trastuzumab emtansine (T-DM1; n = 743) or to trastuzumab (n = 743). Invasive disease-free survival was significantly higher in the T-DM1 group than in the trastuzumab arm (hazard ratio, 0.50; 95% CI, 0.39 to 0.64; P < .001), and risk of distant recurrence was lower in patients who received T-DM1 than in patients who received trastuzumab (hazard ratio, 0.60; 95% CI, 0.45 to 0.79). Grade 3 or higher adverse events occurred in 190 patients (25.7%) who received T-DM1 and in 111 patients (15.4%) who received trastuzumab. RECOMMENDATIONS: Patients with HER2-positive breast cancer with pathologic invasive residual disease at surgery after standard preoperative chemotherapy and HER2-targeted therapy should be offered 14 cycles of adjuvant T-DM1, unless there is disease recurrence or unmanageable toxicity. Clinicians may offer any of the available and approved formulations of trastuzumab, including trastuzumab, trastuzumab and hyaluronidase-oysk, and available biosimilars.Additional information can be found at www.asco.org/breast-cancer-guidelines.

Mite Burden and Immunophenotypic Response to Demodex musculi in Swiss Webster, BALB/c, C57BL/6, and NSG Mice.

Detection methods for Demodex musculi were historically unreliable, and testing was rarely performed because its prevalence in laboratory mice was underestimated. Although infestations are unapparent in most mouse strains, D. musculi burdens are higher and clinical signs detected in various immunodeficient strains. The parasite's influence on the immune system of immunocompetent mice is unknown. We characterized mite burden (immunocompetent and immunodeficient strains) and immunologic changes (immunocompetent strains only) in naïve Swiss Webster (SW; outbred), C57BL/6NCrl (B6; Th1 responder), BALB/cAnNCrl (BALB/c; Th2 responder) and NOD. Cg-Prkdcscid Il2rgtm1Wjl/SzJ (NSG; immunodeficient) mice after exposure to Demodex-infested NSG mice. Infested and uninfested age-matched mice of each strain (n = 5) were euthanized 14, 28, 56, and 112 d after exposure. Mite burden was determined through PCR analysis and skin histopathology; B-cell and CD4+ and CD8+ T-cell counts and activation states (CD25 and CD69) were evaluated by using flow cytometry; CBC counts were performed; and serum IgE levels were measured by ELISA. Mite burden and PCR copy number correlated in NSG mice, which had the highest mite burden, but not in immunocompetent strains. Infested immunocompetent animals developed diffuse alopecia by day 112, and both BALB/c and C57BL/6 mice had significantly increased IgE levels. These findings aligned with the skewed Th1 or Th2 immunophenotype of each strain. BALB/c mice mounted the most effective host response, resulting in the lowest mite burden of all immunocompetent strains at 112 d after infestation without treatment. Clinically significant hematologic abnormalities were absent and immunophenotype was unaltered in immunocompetent animals. Topical treat- ment with imidacloprid-moxidectin (weekly for 8 wk) was effective at eradicating mites by early as 7 d after treatment. IgE levels decreased substantially in infested BALB/c mice after treatment. These findings demonstrate a need for D. musculi surveillance in mouse colonies, because the infestation may influence the use of infested mice in select studies.

Sentinel Lymph Node Biopsy for Patients With Early-Stage Breast Cancer: American Society of Clinical Oncology Clinical Practice Guideline Update.

Purpose To provide current recommendations on the use of sentinel node biopsy (SNB) for patients with early-stage breast cancer. Methods PubMed and the Cochrane Library were searched for randomized controlled trials, systematic reviews, meta-analyses, and clinical practice guidelines from 2012 through July 2016. An Update Panel reviewed the identified abstracts. Results Of the eight publications identified and reviewed, none prompted a change in the 2014 recommendations, which are reaffirmed by the updated literature review. Conclusion Women without sentinel lymph node (SLN) metastases should not receive axillary lymph node dissection (ALND). Women with one to two metastatic SLNs who are planning to undergo breast-conserving surgery with whole-breast radiotherapy should not undergo ALND (in most cases). Women with SLN metastases who will undergo mastectomy should be offered ALND. These three recommendations are based on randomized controlled trials. Women with operable breast cancer and multicentric tumors, with ductal carcinoma in situ, who will undergo mastectomy, who previously underwent breast and/or axillary surgery, or who received preoperative/neoadjuvant systemic therapy may be offered SNB. Women who have large or locally advanced invasive breast cancer (tumor size T3/T4), inflammatory breast cancer, or ductal carcinoma in situ (when breast-conserving surgery is planned) or are pregnant should not undergo SNB.

Recommendations for the Use of WBC Growth Factors: American Society of Clinical Oncology Clinical Practice Guideline Update.

PURPOSE: To update the 2006 American Society of Clinical Oncology guideline on the use of hematopoietic colony-stimulating factors (CSFs). METHODS: The American Society of Clinical Oncology convened an Update Committee and conducted a systematic review of randomized clinical trials, meta-analyses, and systematic reviews from October 2005 through September 2014. Guideline recommendations were based on the review of the evidence by the Update Committee. RESULTS: Changes to previous recommendations include the addition of tbo-filgrastim and filgrastim-sndz, moderation of the recommendation regarding routine use of CSFs in older patients with diffuse aggressive lymphoma, and addition of recommendations against routine dose-dense chemotherapy in lymphoma and in favor of high-dose-intensity chemotherapy in urothelial cancer. The Update Committee did not address recommendations regarding use of CSFs in acute myeloid leukemia or myelodysplastic syndromes in adults. RECOMMENDATIONS: Prophylactic use of CSFs to reduce the risk of febrile neutropenia is warranted when the risk of febrile neutropenia is approximately 20% or higher and no other equally effective and safe regimen that does not require CSFs is available. Primary prophylaxis is recommended for the prevention of febrile neutropenia in patients who are at high risk on the basis of age, medical history, disease characteristics, and myelotoxicity of the chemotherapy regimen. Dose-dense regimens that require CSFs should only be used within an appropriately designed clinical trial or if supported by convincing efficacy data. Current recommendations for the management of patients exposed to lethal doses of total-body radiotherapy, but not doses high enough to lead to certain death as a result of injury to other organs, include the prompt administration of CSFs.

A comparison of mouse parvovirus 1 infection in BALB/c and C57BL/6 mice: susceptibility, replication, shedding, and seroconversion.

This study characterized the effects of challenge with a field isolate of mouse parvovirus 1 (MPV1e) in C57BL/6NCrl (B6) and BALB/cAnNCrl (C) mice. We found that C mice were more susceptible to MPV1e infection than were B6 mice; ID50 were 50 to 100 times higher after gavage and 10-fold higher after intraperitoneal injection in B6 as compared with C mice. To evaluate the host strain effect on the pathogenesis of MPV1e, B6 and C mice were inoculated by gavage. Feces and tissues, including mesenteric lymph nodes (MLN), ileum, spleen and blood, were collected for analysis by quantitative PCR (qPCR) to assess infection and fecal shedding and by RT-qPCR to evaluate replication. Peak levels of MPV1e shedding, infection, and replication were on average 3.4, 4.3, and 6.2 times higher, respectively, in C than in B6 mice. Peaks occurred between 3 and 10 d after inoculation in C mice but between 5 and 14 d in B6 mice. Multiplexed fluorometric immunoassays detected seroconversion in 2 of 3 C mice at 7 d after inoculation and in all 3 B6 mice at 10 d. By 56 d after inoculation, viral replication was no longer detectable, and fecal shedding was very low; infection persisted in ileum, spleen, and MLN, with levels higher in C than B6 mice and highest in MLN. Therefore, the lower susceptibility of B6 mice, as compared with C mice, to MPV1e infection was associated with lower levels of infection, replication, and shedding and delayed seroconversion.

Sentinel lymph node biopsy for patients with early-stage breast cancer: American Society of Clinical Oncology clinical practice guideline update.

PURPOSE: To provide evidence-based recommendations to practicing oncologists, surgeons, and radiation therapy clinicians to update the 2005 clinical practice guideline on the use of sentinel node biopsy (SNB) for patients with early-stage breast cancer. METHODS: The American Society of Clinical Oncology convened an Update Committee of experts in medical oncology, pathology, radiation oncology, surgical oncology, guideline implementation, and advocacy. A systematic review of the literature was conducted from February 2004 to January 2013 in Medline. Guideline recommendations were based on the review of the evidence by Update Committee. RESULTS: This guideline update reflects changes in practice since the 2005 guideline. Nine randomized clinical trials (RCTs) met systematic review criteria for clinical questions 1 and 2; 13 cohort studies informed clinical question 3. RECOMMENDATIONS: Women without sentinel lymph node (SLN) metastases should not receive axillary lymph node dissection (ALND). Women with one to two metastatic SLNs planning to undergo breast-conserving surgery with whole-breast radiotherapy should not undergo ALND (in most cases). Women with SLN metastases who will undergo mastectomy should be offered ALND. These three recommendation are based on RCTs. Women with operable breast cancer and multicentric tumors, with ductal carcinoma in situ (DCIS) who will undergo mastectomy, who previously underwent breast and/or axillary surgery, or who received preoperative/neoadjuvant systemic therapy may be offered SNB. Women who have large or locally advanced invasive breast cancer (tumor size T3/T4), inflammatory breast cancer, or DCIS (when breast-conserving surgery is planned) or are pregnant should not undergo SNB. These recommendations are based on cohort studies and/or informal consensus. In some cases, updated evidence was insufficient to update previous recommendations.

Efficacy of direct detection of pathogens in naturally infected mice by using a high-density PCR array.

We used a high-density array of real-time PCR assays for commonly reported rodent infectious agents (PRIA) to test naturally infected index mice and sentinel mice exposed by contact and soiled-bedding transfer. PRIA detected 14 pathogens--including viruses, bacteria, fur mites, pinworms, and enteric protozoa--in 97.2% of 28 pooled fecal samples, fur-perianal swabs, and oral swabs from 4 cages containing a total of 10 index mice. Among these pathogens, PRIA (like conventional health monitoring methods) failed to detect Mycoplasma pulmonis, Pasteurella pneumotropica, and Giardia spp. in all of the 9 contact and 9 soiled-bedding sentinels. PRIA demonstrated murine adenovirus and Cryptosporidium and Spironucleus spp. in contact but not soiled-bedding sentinels and detected Helicobacter and pinworms in fewer than half of the soiled-bedding sentinels. Of the 4 species of Helicobacter that species-specific PCR assays identified in index mice, only H. ganmani was found in soiled-bedding and contact sentinels. PRIA detected all of the pathogens in sentinels that were identified by conventional methods. Myobia musculi was detected by PCR in index and sentinel mice but missed by conventional parasitologic examinations. In summary, PRIA reproducibly detected diverse pathogens in heavily pooled specimens collected noninvasively from infected index mice antemortem. The inability of PRIA and conventional health monitoring methods (that is, parasitology, micro-biology, and serology) to demonstrate transmission of some pathogens to contact sentinels and the inefficient transmission of others to soiled-bedding sentinels underscores the importance of direct PCR testing to determine the pathogen status of rodents in quarantine and during routine colony surveillance.

Evaluation of diagnostic methods for Myocoptes musculinus according to age and treatment status of mice (Mus musculus).

Detecting and controlling murine fur mites continues to be challenging. Here we compared the efficacy of fur-pluck, cage PCR, and fur PCR testing of mice naturally infested with Myocoptes musculinus and make recommendations regarding the application of these diagnostic strategies in aged or treated mice. We compared all 3 diagnostic methods in groups of infested and noninfested control mice over time. For fur plucks, we used a scoring system to quantitatively compare mite infestations across ages. Mice that were 4 wk old had higher egg and mite scores than did older mice, with average scores at 4 wk corresponding to 40 to 100 individual fur mites and eggs per sample. Furthermore, 15% and 20% of samples from infested mice at 24 and 28 wk of age, respectively, lacked all fur mites and eggs. Cage PCR results varied as mice grew older. Fur PCR testing was the most sensitive and specific assay in untreated infested mice, particularly when mite densities were low. In addition, we compared fur-pluck and fur PCR tests for evaluating the efficacy of selamectin treatment. Two treatments with selamectin eliminated Myocoptes fur-mite infestations. At 8 wk after treatment, all fur-pluck samples were negative, but one-third of treated infested cages remained positive by fur PCR assay; at 16 wk after treatment, all cages were negative by fur PCR assay. Because offspring of infested mice were invariably heavily infested, breeding of suspected infested mice with subsequent testing of offspring was the definitive testing strategy when fur-pluck and PCR results conflicted.

Multiple effects of dendritic cell depletion on murine norovirus infection.

Dendritic cells (DCs) are permissive to murine norovirus (MNV) infection in vitro and in vivo. However, their roles during infection in vivo are not well defined. To determine the role of DCs during infection, conventional DCs were depleted from CD11c-DTR mice and infected with a persistent MNV strain. Viral titres in the intestine and secondary lymphoid organs were determined at early time points during infection, and anti-MNV antibody responses were analysed later during infection. Depletion of conventional DCs resulted in increased viral loads in intestinal tissues, impaired generation of antibody responses, and a failure of MNV to efficiently infect lymphoid tissues. These data suggest that DCs play multiple roles in MNV pathogenesis, in both innate immunity and the efficient generation of adaptive immune responses against MNV, as well as by promoting the dissemination of MNV to secondary lymphoid tissues. This is the first study to probe the roles of DCs in controlling and/or facilitating a norovirus infection in vivo and provides the basis for further studies aimed at defining mechanisms by which DCs control MNV replication and promote viral dissemination.

Murine noroviruses bind glycolipid and glycoprotein attachment receptors in a strain-dependent manner.

Human norovirus infections are the most common cause of acute nonbacterial gastroenteritis in humans worldwide, and glycan binding plays an important role in the susceptibility to these infections. However, due to the lack of an efficient cell culture system or small animal model for human noroviruses, little is known about the biological role of glycan binding during infection. Murine noroviruses (MNV) are also enteric viruses that bind to cell surface glycans, but in contrast to their human counterparts, they can be grown in tissue culture and a small animal host. In this study, we determined glycan-binding specificities of the MNV strains MNV-1 and CR3 in vitro, identified molecular determinants of glycan binding, and analyzed infection in vivo. We showed that unlike MNV-1, CR3 binding to murine macrophages was resistant to neuraminidase treatment and glycosphingolipid depletion. Both strains depended on N-linked glycoproteins for binding, while only MNV-1 attachment to macrophages was sensitive to O-linked glycoprotein depletion. In vivo, CR3 showed differences in tissue tropism compared to MNV-1 by replicating in the large intestine. Mapping of a glycan-binding site in the MNV-1 capsid by reverse genetics identified a region topologically similar to the histo-blood group antigen (HBGA)-binding sites of the human norovirus strain VA387. The recombinant virus showed distinct changes in tissue tropism compared to wild-type virus. Taken together, our data demonstrate that MNV strains evolved multiple strategies to bind different glycan receptors on the surface of murine macrophages and that glycan binding contributes to tissue tropism in vivo.

Why current breast pathology practices must be evaluated. A Susan G. Komen for the Cure white paper: June 2006.

Ending breast cancer has long been the mission of Susan G. Komen for the Cure. To this end, the organization has a strong interest and proven track record in ensuring public investment in quality breast health and breast cancer care. Recently, Susan G. Komen for the Cure identified major issues in the practice of pathology that have a negative impact on the lives of thousands of breast cancer patients in the United States. These issues were identified through a comprehensive literature review and interviews conducted in 2005-2006 with experts in oncology, breast pathology, surgery, and radiology. The interviewees practiced in community, academic, and cooperative group settings. Komen for the Cure has identified four areas that have a direct impact on the quality of care breast cancer patients receive in the United States, the accuracy of breast pathology diagnostics, the effects of current health insurance, and reimbursement policies on patients who are evaluated for a possible breast cancer diagnosis, the substantial decrease in tissue banking participation, particularly during a time of rapid advances in biologically correlated clinical science and the role for the Susan G. Komen for the Cure, pathology professional societies and the Federal government in ensuring that breast pathology practices meet the highest possible standards in the United States Concerns surrounding the quality and practice of breast pathology are not limited to diagnostic accuracy. Other considerations include, training and proficiency of pathologists who are evaluating breast specimens, the lack of integration of pathologists in the clinical care team, inadequate compensation for the amount of work required to thoroughly analyze specimens, potential loss in translational research as a result of medical privacy regulations, and the lack of mandatory uniform pathology practice standards without any way to measure the degree of variation or to remedy it.

Breast cancer patients' perspectives on and use of complementary and alternative medicine: a study by the Susan G. Komen Breast Cancer Foundation.

The purpose of this study was to examine patterns and predictors of complementary and alternative medicine (CAM) among breast cancer patients. A review of the existing survey literature on CAM use for breast cancer was conducted with a series of eight focus groups (N = 67) to further examine the perspectives of breast cancer patients on CAM. The rates of CAM use varied from 17 to 75%, with a mean of 45%. Vitamins and minerals and herbs were the most frequently cited categories. Users tended to be younger, more educated, and more likely to have used CAM prior to their diagnosis. Focus group data indicate that breast cancer patients use a wide array of CAM for a variety of reasons, including symptom management, improving quality of life, and enhancing immune function. Although women rely on a variety of resources for information, they frequently experience frustration owing to the absence or conflicting nature of such information. Communication with conventional providers about CAM is frequently experienced as either unsupportive or not helpful by many patients. The results point to the value of developing better evidence-based informational resources related to CAM and cancer and the need for physicians to become better educated about CAM and how to communicate more effectively with their breast cancer patients about it.