Clinical standards for the assessment, management and rehabilitation of post-TB lung disease.

BACKGROUND: Increasing evidence suggests that post-TB lung disease (PTLD) causes significant morbidity and mortality. The aim of these clinical standards is to provide guidance on the assessment and management of PTLD and the implementation of pulmonary rehabilitation (PR).METHODS: A panel of global experts in the field of TB care and PR was identified; 62 participated in a Delphi process. A 5-point Likert scale was used to score the initial ideas for standards and after several rounds of revision the document was approved (with 100% agreement).RESULTS: Five clinical standards were defined: Standard 1, to assess patients at the end of TB treatment for PTLD (with adaptation for children and specific settings/situations); Standard 2, to identify patients with PTLD for PR; Standard 3, tailoring the PR programme to patient needs and the local setting; Standard 4, to evaluate the effectiveness of PR; and Standard 5, to conduct education and counselling. Standard 6 addresses public health aspects of PTLD and outcomes due to PR.CONCLUSION: This is the first consensus-based set of Clinical Standards for PTLD. Our aim is to improve patient care and quality of life by guiding clinicians, programme managers and public health officers in planning and implementing adequate measures to assess and manage PTLD.

Quantitative proteomic comparison of salt stress in Chlamydomonas reinhardtii and the snow alga Chlamydomonas nivalis reveals mechanisms for salt-triggered fatty acid accumulation via reallocation of carbon resources.

BACKGROUND: Chlamydomonas reinhardtii is a model green alga strain for molecular studies; its fully sequenced genome has enabled omic-based analyses that have been applied to better understand its metabolic responses to stress. Here, we characterised physiological and proteomic changes between a low-starch C. reinhardtii strain and the snow alga Chlamydomonas nivalis, to reveal insights into their contrasting responses to salinity stress. RESULTS: Each strain was grown in conditions tailored to their growth requirements to encourage maximal fatty acid (as a proxy measure of lipid) production, with internal controls to allow comparison points. In 0.2 M NaCl, C. nivalis accumulates carbohydrates up to 10.4% DCW at 80 h, and fatty acids up to 52.0% dry cell weight (DCW) over 12 days, however, C. reinhardtii does not show fatty acid accumulation over time, and shows limited carbohydrate accumulation up to 5.5% DCW. Analysis of the C. nivalis fatty acid profiles showed that salt stress improved the biofuel qualities over time. Photosynthesis and respiration rates are reduced in C. reinhardtii relative to C. nivalis in response to 0.2 M NaCl. De novo sequencing and homology matching was used in conjunction with iTRAQ-based quantitative analysis to identify and relatively quantify proteomic alterations in cells exposed to salt stress. There were abundance differences in proteins associated with stress, photosynthesis, carbohydrate and lipid metabolism proteins. In terms of lipid synthesis, salt stress induced an increase in dihydrolipoyl dehydrogenase in C. nivalis (1.1-fold change), whilst levels in C. reinhardtii remained unaffected; this enzyme is involved in acetyl CoA production and has been linked to TAG accumulation in microalgae. In salt-stressed C. nivalis there were decreases in the abundance of UDP-sulfoquinovose (- 1.77-fold change), which is involved in sulfoquinovosyl diacylglycerol metabolism, and in citrate synthase (- 2.7-fold change), also involved in the TCA cycle. Decreases in these enzymes have been shown to lead to increased TAG production as fatty acid biosynthesis is favoured. Data are available via ProteomeXchange with identifier PXD018148. CONCLUSIONS: These differences in protein abundance have given greater understanding of the mechanism by which salt stress promotes fatty acid accumulation in the un-sequenced microalga C. nivalis as it switches to a non-growth state, whereas C. reinhardtii does not have this response.

Practices and opinions of New Zealand veterinarians regarding control of bovine viral diarrhoea.

AIMS: To explore recommendations that New Zealand veterinarians make for diagnosing and managing bovine viral diarrhoea (BVD) in cattle herds under different clinical scenarios and their opinions towards potential barriers and opportunities for implementing BVD control programmes in New Zealand. METHODS: A cross-sectional survey of registered veterinarians in New Zealand was conducted in 2019. Respondents were asked about the approaches they would use to manage BVD under different clinical scenarios as well as their opinions on national BVD control. A subset of veterinarians completed a more in-depth survey providing additional free-text responses on a range of different BVD topics. Descriptive statistics were provided for all quantitative study variables and the free-text responses were also analysed to generate further insights into veterinarians' perceptions towards BVD management. RESULTS: The cross-sectional survey was completed by 101 of an estimated 870 (11.6%) cattle veterinarians. Thirty-five veterinarians completed the in-depth survey. There was wide variation in the BVD diagnostic testing and vaccination protocols that respondents recommended under different clinical scenarios. Annual bulk milk BVD testing was perceived as a valuable tool for initiating BVD discussions with dairy farmers. Respondents indicated that beef farmers were more difficult to engage in BVD control largely due to the logistical challenges of yarding cattle at the appropriate times to implement interventions, with many farmers only contacting veterinarians after experiencing a BVD outbreak Most respondents (91/101; 90%) believed it was possible to eradicate BVD from New Zealand, but cited lack of farmer awareness and poor compliance with management recommendations as significant barriers. The measure with the most support for inclusion in a compulsory national eradication programme was requiring farmers to declare the status of their animals prior to sale while the least supported measure was requiring farmers to double fence boundaries to prevent nose-to-nose contact with neighbouring stock. Although respondents highlighted the need for farmers and industry to support any national eradication programme in order for it to be successful, there was also recognition that veterinarians could be more pro-active in engaging with farmers particularly in discussions around the economics of BVD. CONCLUSIONS AND CLINICAL RELEVANCE: While the survey respondents appeared to be highly supportive of BVD control, it was perceived that financial and logistical barriers existed that could impede farmer engagement. Further extension efforts may be needed to ensure that veterinarians are presenting clear and consistent recommendations about BVD management to farmers.Abbreviations: BVD: Bovine viral diarrhoea; NAIT: National Animal Identification and Tracing System; PI: Persistently infected.

Effects of Racism on Oral Health in the United States.

This paper comments on the Effects of Racism on Oral Health in the United States (US). It provides the background and sets the stage to raise questions about race: how was race defined originally, what exactly is race, and how have racial categories been enumerated? Following this path, the paper broadens the scope of view regarding data attributable to racial categories pointing to social and cultural factors that influence overall health outcomes, particularly those related to oral health. Oral health researchers, advocates, providers, administrators, program planners, and funders, among others rely on data, often compiled by racial categories. We should be aware of potential vagaries that can accompany race-based data, and its interpretation and application, regarding oral health. The paper suggests we should be mindful of other influences that affect documented differences among populations regarding their oral health status.

Temporal trends in bulk tank milk antibody ELISA and PCR test results for bovine viral diarrhoea in New Zealand pastoral dairy herds.

Aims: To describe temporal trends in bulk milk antibody ELISA and PCR testing for bovine viral diarrhoea (BVD) in New Zealand pastoral dairy herds and to assess the use of historical accession data to predict herd-level BVD incursions. Methods: Data on all diagnostic testing of bulk milk for BVD performed by the Livestock Improvement Corporation (Hamilton, NZ) over eight lactation seasons from 1 June 2010 to 31 May 2018 were analysed. This included anonymised herd identification, geographic location, herd size, sample collection date, sample to positive (S/P) ratio for antibody ELISA results, and cycle threshold values for PCR detecting viral RNA. Multivariable logistic regression was used to explore the relationship between historical accession data and the risk of herds having at least one positive bulk milk PCR test result in the 2017 season. Results: There were 156,034 bulk milk BVD diagnostic testing accessions for 10,495 uniquely identified dairy herds over the 8-season period. The prevalence of tested herds with at least one positive bulk milk PCR test result decreased from 14.6% (407/2,786) in the 2010 season to 5.6% (355/6,309) in the 2017 season with similarly marked declines in S/P ratios. In the 2017 season, 2,961/6,309 (46.9%) herds had S/P ratios greater than the 0.75 cut-off value indicating recent or active BVD virus transmission within the herd while 1,422/6,309 (22.5%) herds were classified as having negative or low S/P ratios. Herds that cleared BVD from the milking herd experienced a mean decline in S/P ratio of 0.11 units per year (min 0.05; max 0.18). In the multivariable analysis, the overall incidence risk of herds experiencing a BVD incursion in the 2017 season was 3.8% (146/3,848) and there were three significant predictors in the final model: herd size, PCR status in the 2014 season, and change in S/P ratio between the 2014 and 2015 seasons. The area under the receiver operating curve for the final model was 0.695 indicating poor discrimination. Conclusions and clinical relevance: The prevalence of dairy herds in New Zealand with positive bulk milk PCR test results and high S/P ratios has decreased over time, suggesting fewer herds are actively infected with BVD and that herd immunity may also be declining. Although monitoring trends in bulk milk test results provides useful information on changes in individual herd status, it is difficult to accurately predict when new incursions will occur and farmers should continue to maintain good biosecurity.

Post-tuberculosis lung health: perspectives from the First International Symposium.

ALTHOUGH CURABLE, TB frequently leaves the individual with chronic physical and psycho-social impairment, but these consequences have been largely neglected. The 1st International Post-Tuberculosis Symposium (Stellenbosch, South Africa) was held to discuss priorities and gaps in addressing this issue. A barrier to progress has been the varied terminology and nomenclature, so the Delphi process was used to achieve consensus on definitions. Lack of sufficient evidence hampered definitive recommendations in most domains, including prevention and treatment of post-TB lung disease (PTLD), but the discussions clarified the research needed. A consensus was reached on a toolkit for future PTLD measurement and on PTLD patterns to be considered. The importance of extra-pulmonary consequences and progressive impairment throughout the life-course was identified, including TB recurrence and increased mortality. Patient advocates emphasised the need to address the psychological and social impacts post TB and called for clinical guidance. More generally, there is an urgent need for increased awareness and research into post-TB complications.

Serological evidence for exposure to bovine viral diarrhoea virus in sheep co-grazed with beef cattle in New Zealand.

Aims: To determine whether sheep that co-grazed with cattle that were suspected to be positive for bovine viral diarrhoea (BVD) virus had serological evidence of exposure to the virus.Methods: Eighteen commercial farms that routinely co-grazed cattle and sheep in the same paddocks were recruited through purposive sampling. The recruiting veterinarians identified nine farms with cattle herds that were known or highly suspected to be positive for BVD and nine farms that were considered to be free of BVD. Blood samples were taken from 15 ewes aged 1 year on each farm and samples were submitted to a commercial diagnostic laboratory to test for antibodies against pestiviruses using an ELISA. All samples that were positive were then tested using a virus neutralisation test (VNT)for antibodies against BVD virus.Results: Of the 270 blood samples, 17 were positive for pestivirus antibodies by ELISA and these originated from two farms that were known or suspected to have BVD virus-positive cattle. None of the samples from the nine flocks co-grazed with cattle herds that were known or suspected to be BVD virus-negative were positive for pestivirus antibodies. Within the two positive farms, 2/15 samples from the first farm and 15/15 samples from the second farm were antibody-positive. When the 17 positive blood samples were submitted for VNT, all 15 samples from the second farm tested positive for BVD virus antibodies with the highest titre being 1:512.Conclusions and clinical relevance: In this small sample of New Zealand sheep and beef farms with suspected BVD infection in cattle, there was evidence of pestivirus exposure in co-grazed sheep. Although we were unable to confirm the origin of the exposure in these sheep, these findings highlight that farmers who are trying to eradicate BVD from their cattle should be mindful that the infection may also be circulating in sheep, and both populations should be considered a possible risk to each other for generating transient and persistent infections. Further work is needed to estimate the true prevalence of New Zealand sheep flocks that are affected by BVD and the associated economic impacts.

Practices and opinions of New Zealand beef cattle farmers towards bovine viral diarrhoea control in relation to real and perceived herd serological status.

Aims: To investigate the seroprevalence of infection with bovine viral diarrhoea (BVD) virus among 75 beef herds and seroconversion in cattle during early pregnancy, and to determine the practices and opinions of farmers towards BVD control and their association with real and perceived herd serological status.Methods: Blood samples were collected before mating in 75 beef herds across New Zealand from 15 unvaccinated heifers that had delivered their first calf that season. Serum samples were tested for BVD antibodies using ELISA individually, and after pooling samples for each farm. Animals that were antibody-negative were retested at either pregnancy diagnosis or weaning. Farmers were asked to complete a detailed survey about herd demographics, BVD testing and vaccination practices, and opinions towards national BVD control.Results: Based on the pooled serum antibody ELISA results, there were 28/75 (37%) negative herds, 15/75 (20%) suspect herds, and 32/75 (43%) positive herds. Of 1,117 animals sampled 729 (65.3%) tested negative for BVD virus antibodies; when retested, 47/589 (8.0%) animals from 13/55 (24%) herds had seroconverted. Among 71 famers providing survey responses 11 (15%) believed their herd was infected with BVD, 24 (34%) were unsure and 36 (51%) did not think their herd was infected. Only 19/71 (18%) farmers had performed any BVD testing within the past 5 years and 50/70 (71%) had not vaccinated any cattle for BVD. Support for national BVD eradication programme was strong in 51/71 (56%) respondents, but the biggest challenge to BVD control was considered to be famer compliance. Compared to farmers who did not think their herd was infected, more farmers who thought BVD was present in their herds had previously tested for BVD, would consider testing all replacement calves, and would support establishing a national BVD database; fewer would consider purchasing BVD tested or vaccinated cattle only.Conclusions and clinical relevance: Only 15% of the beef farmers in this study believed their herds were infected with BVD virus and few of them had undertaken BVD screening. Nevertheless many were supportive of implementing a national BVD control programme. It is likely that the lack of farmer awareness around BVD and the failure of farmers to recognise the potential impacts in their herds are hindering progress in controlling the disease in New Zealand. There are opportunities for New Zealand veterinarians to be more proactive in helping beef farmers explore BVD management options.

Recommendations for the testing and control of bovine viral diarrhoea in New Zealand pastoral cattle production systems.

Eradicating bovine viral diarrhoea (BVD) from cattle populations requires a clear approach for determining the epidemiological status of individual herds and implementing the appropriate control measures to ensure the transmission cycle is cost-effectively broken. This is particularly important in countries such as New Zealand where there is currently no coordinated national programme and the herd-level decisions to control BVD are left to the discretion of individual farmers and veterinarians. To ensure greater consistency in the information being delivered by different stakeholders, we review the epidemiology of BVD in the context of New Zealand pastoral production systems and provides a series of simplified recommendations for the future control of BVD in beef and dairy herds. Based on analysis of BVD test accession data from commercial diagnostic laboratories, it has been estimated that 40.6% of dairy herds and 45.6% of beef herds tested had positive results for antibodies to BVD virus. While BVD continues to remain widespread and under voluntary control in New Zealand, it is recommended that herds test all individual mixed-age cows and replacement heifers for BVD virus or antigen and remove persistently infected animals from the breeding population. All new breeding animals that have entered the herd either through purchase or birth should also be tested for BVD virus. Biosecurity risks should be managed by reducing contacts with other herds and implementing targeted vaccination programmes. All individual purchased cattle should be tested and confirmed negative for BVD virus before being moved onto the buyer's property, even if the herd of origin had a negative antibody-based screening test. Herds should continue annual antigen or virus testing of all calves as soon as possible after birth to identify any persistently infected animals.

Assessing the use of diagnostic laboratory accession data to support national bovine viral diarrhoea control in New Zealand.

Aims: To assess the suitability of using existing national diagnostic laboratory testing data to support national bovine viral diarrhoea (BVD) research, surveillance, and control in New Zealand. Methods: Data on laboratory accessions for BVD diagnostic testing in New Zealand from 1 January 2015 to 31 December 2017 were provided by four commercial veterinary diagnostic companies. The data were integrated into a single dataset containing the unique accession number, sample submission date, farm location (territorial authority level), test type (bulk milk antibody-ELISA, bulk milk PCR, serum antibody-ELISA, blood/serum/tissue antigen-ELISA, or blood/serum/tissue PCR), and test results. Estimates for the number of registered cattle farms in each territorial authority were generated from the National Animal Identification and Tracing database. Results were summarised for July 2015 to June 2016 and July 2016 to June 2017. Results: There was a total of 59,007 unique BVD diagnostic test accessions including 39,920 (67.6%) for bulk milk antibody-ELISA, 27,832 (47.2%) for bulk milk PCR, 3,229 (5.5%) for serum antibody-ELISA, 9,132 (15.5%) for blood/serum/tissue antigen-ELISA, and 7,122 (12.1%) for blood/serum/tissue PCR. Of the 17,946 accessions for blood/serum/tissue samples, 4,316 (24.0%) were missing the herd production type and 6,678 (37.2%) were missing the animals age. Approximately 7,000/10,958 (65%) dairy herds and 1,600/43,611 (4%) beef herds were conducting annual BVD screening tests. In 2016/2017, the prevalence of accessions with ≥1 BVD-positive result was 40.6% for bulk milk antibody, 6.4% for bulk milk PCR, 45.6% for serum antibody, and 9.8% for blood/serum/tissue antigen-ELISA or PCR tests. There was substantial regional variation in both the percentage of herds testing for BVD and the prevalence of positive accessions. Following pooled serum antibody-ELISA, only 175/604 (29.0%) beef herds and 177/566 (31.3%) dairy herds had recorded follow-up testing. Conclusions and Clinical Relevance: Laboratory diagnostic accession data has the potential to provide valuable insights about BVD epidemiology in New Zealand, but there are significant limitations in the data collected and discrepancies in the different systems that each laboratory uses to measure, interpret, and record diagnostic data. There is a strong need to develop a more consistent national system for recording and sharing BVD test results to support BVD management at farm and industry levels. Abbreviations: BVD: Bovine viral diarrhoea; Ct: Cycle threshold; NAIT: National Animal Identification and Tracing; NZVP: New Zealand Veterinary Pathology; PI: Persistently infected; S/P: Sample to positive control.

Probiotics in Goat Milk Products: Delivery Capacity and Ability to Improve Sensory Attributes.

Dairy foods, particularly those of bovine origin, are the predominant vehicles for delivery of probiotic bacteria. Caprine (goat) milk also possesses potential for successful delivery of probiotics, and despite its less appealing flavor in some products, the use of goat milk as a probiotic carrier has rapidly increased over the last decade. This review reports on the diversity, applicability, and potential of using probiotics to enhance the sensory properties of goat milk and goat milk-based products. A brief conceptual introduction to probiotic microorganisms is followed by an account of the unique physicochemical, nutritive, and beneficial aspects of goat milk, emphasizing its advantages as a probiotic carrier. The sensory properties of probiotic-enriched goat milk products are also discussed. The maintenance of probiotic viability and desirable physicochemical characteristics in goat milk products over shelf life is possible. However, the unpleasant sensory features of some goat milk products remain a major disadvantage that hinder its wider utilization. Nevertheless, certain measures such as fortification with selected probiotic strains, inclusion of fruit pulps and popular flavor compounds, and production of commonly consumed tailor-made goat milk-based products have potential to overcome this limitation. In particular, certain probiotic bacteria release volatile compounds as a result of their metabolism, which are known to play a major role in the aroma profile and sensory aspects of the final products.

Seroprevalence of antibodies to Pestivirus infections in South Australian sheep flocks.

OBJECTIVE: Bovine viral diarrhoea virus (BVDV) and border disease virus (BDV) are of the genus Pestivirus. They are known to cause significant reproductive and production losses, with BVDV acknowledged as a major source of economic loss to the Australian cattle industry. Very little is currently known about the prevalence and effect of pestiviruses in the Australian sheep industry. The present study aimed to examine the seroprevalence and effect of both BVDV and BDV in South Australian sheep flocks. METHODS: In total, 875 breeding ewes on 29 properties were serologically tested by ELISA, AGID and VNT assays for the presence of Pestivirus-specific antibodies. RESULTS: Three (0.34%) individual animals returned serological results suggestive of previous BDV infection. All three positive animals were collected from one property, giving a property level seroprevalence of 3.45% and a within-flock seroprevalence of 10%. CONCLUSION: The results suggested that BDV infection is present, albeit at a very low incidence, in the South Australian sheep flock and BVDV infection appears to be absent. Consequently, pestiviruses are unlikely to impair production in South Australian sheep populations.

BVDV in Australian alpacas: natural infection and clinical profiles following co-mingling with a persistently infected heifer.

BACKGROUND: Although predominantly a disease of cattle, bovine viral diarrhoea virus (BVDV) is known to infect other ruminant and camelid species such as sheep and alpacas. The aims of this study were to determine if BVDV-naive alpacas would become acutely infected and seroconvert to the predominant Australian strain of BVDV following co-mingling with a BVDV-1c persistently infected (PI) heifer and to determine what, if any, clinical signs, haematological responses and selected biochemical changes occur with acute BVDV-1c infections in alpacas. METHODS: A PI heifer and four alpacas co-mingled for 2 weeks. Weekly blood samples were collected and twice weekly clinical examinations were performed on the alpacas. RESULTS: Serum analysis by antibody ELISA indicated that all four alpacas were positive for BVDV-specific antibodies between 35 and 54 days after mixing with the BVDV-1c PI heifer. Viral antigen was detected by antigen ELISA in two alpacas on days 21 and 35 after initial mixing. In general, all the physical clinical parameters measured were normal. Serum biochemical and haematological analyses in two of the alpacas revealed marginally low sodium, chloride and elevated potassium concentrations, a lymphocytosis, monocytosis and a neutrophilia at some point during the study period in either one or both of the alpacas. CONCLUSION: This study showed that infection in Australian alpacas readily occurs when a BVDV-1c PI bovine co-mingles with naive alpacas and that acute infections are clinically mild and undetectable without serological testing.

Potential influence of dairy propionibacteria on the growth and acid metabolism of Streptococcus bovis and Megasphaera elsdenii.

Ruminal acidosis is a prevalent disorder among dairy cows and feedlot cattle, which can significantly impair their health and productivity. This study, involving seven different strains of dairy propionibacteria, represents an in vitro investigation of the feasibility of using these organisms as direct-fed microbials to control lactic acid acumulation in the rumen. Interactions between the propionibacteria, Streptococcus bovis and Megasphaera elsdenii were evaluated in terms of effects on lactic, acetic and propionic acid metabolism, following co-incubation. Spot resistance tests showed slight but varying degrees of growth inhibition by S. bovis among the propionibacteria, while no inhibition was observed between M. elsdenii and the different strains of dairy propionibacteria. In the co-culture experiments comprising S. bovis in nutrient broth, significant differences in pH and the levels of production of lactic, acetic and propionic acid, were observed between treatments following inoculation with various propionibacteria and/or M. elsdenii. In general, lactic acid concentrations at the end of the incubation were significantly lower in the cultures containing propionibacteria compared with cultures comprising either S. bovis only or S. bovis + M. elsdenii, although efficacy of lactate metabolism varied between species and strains. Moreover,the accumulation of acetic and propionic acid in the combined cultures, but not in the solo S. bovis culture, indicated that these compounds were produced as a result of the metabolism of lactic acid by the propionibacteria and M. elsdenii.

Antibodies to bovine viral diarrhoea virus (BVDV) in water buffalo (Bubalus bubalis) and cattle from the Northern Territory of Australia.

BACKGROUND: Farmed and feral water buffaloes (Bubalus bubalis) populations often coexist with cattle in the Northern Territory of Australia, but their level of exposure to bovine viral diarrhoea virus (BVDV) is unknown. METHODS: Water buffalo (n = 245) and cattle (n = 184) serum samples were collected by the NT Government as part of an ongoing disease surveillance scheme at varying intervals between 1993 and 2001. All samples were frozen and stored at -80°C until testing. Water buffalo samples from farming properties were identified as 'farmed' animals and the remaining samples as 'feral' populations. Serum samples were analysed using commercially available ELISAs to test for the presence of BVDV antibodies. RESULTS: Testing of historical water buffalo sera for BVDV antibodies revealed a low level of exposure, with 4.5% (95% CI ± 2.6%) being sero-positive; cattle from the same geographical area and time period had higher levels of exposure at 74.5% (95% CI ± 6.3%). DISCUSSION: This survey showed that water buffalo are susceptible to infection with BVDV. No persistently infected water buffalo were identified in this study.

Oestrogen receptor positive breast cancer metastasis to bone: inhibition by targeting the bone microenvironment in vivo.

Clinical trials have shown that adjuvant Zoledronic acid (ZOL) reduces the development of bone metastases irrespective of ER status. However, post-menopausal patients show anti-tumour benefit with ZOL whereas pre-menopausal patients do not. Here we have developed in vivo models of spontaneous ER+ve breast cancer metastasis to bone and investigated the effects of ZOL and oestrogen on tumour cell dissemination and growth. ER+ve (MCF7, T47D) or ER-ve (MDA-MB-231) cells were administered by inter-mammary or inter-cardiac injection into female nude mice ± estradiol. Mice were administered saline or 100 µg/kg ZOL weekly. Tumour growth, dissemination of tumour cells in blood, bone and bone turnover were monitored by luciferase imaging, histology, flow cytometry, two-photon microscopy, micro-CT and TRAP/P1NP ELISA. Estradiol induced metastasis of ER+ve cells to bone in 80-100 % of animals whereas bone metastases from ER-ve cells were unaffected. Administration of ZOL had no effect on tumour growth in the fat pad but significantly inhibited dissemination of ER+ve tumour cells to bone and frequency of bone metastasis. Estradiol and ZOL increased bone volume via different mechanisms: Estradiol increased activity of bone forming osteoblasts whereas administration of ZOL to estradiol supplemented mice decreased osteoclast activity and returned osteoblast activity to levels comparable to that of saline treated mice. ER-ve cells require increased osteoclast activity to grow in bone whereas ER+ve cells do not. Zol does not affect ER+ve tumour growth in soft tissue, however, inhibition of bone turnover by ZOL reduced dissemination and growth of ER+ve breast cancer cells in bone.

Human breast cancer bone metastasis in vitro and in vivo: a novel 3D model system for studies of tumour cell-bone cell interactions.

Bone is established as the preferred site of breast cancer metastasis. However, the precise mechanisms responsible for this preference remain unidentified. In order to improve outcome for patients with advanced breast cancer and skeletal involvement, we need to better understand how this process is initiated and regulated. As bone metastasis cannot be easily studied in patients, researchers have to date mainly relied on in vivo xenograft models. A major limitation of these is that they do not contain a human bone microenvironment, increasingly considered to be an important component of metastases. In order to address this shortcoming, we have developed a novel humanised bone model, where 1 × 10(5) luciferase-expressing MDA-MB-231 or T47D human breast tumour cells are seeded on viable human subchaodral bone discs in vitro. These discs contain functional osteoclasts 2-weeks after in vitro culture and positive staining for calcine 1-week after culture demonstrating active bone resorption/formation. In vitro inoculation of MDA-MB-231 or T47D cells colonised human bone cores and remained viable for <4 weeks, however, use of matrigel to enhance adhesion or a moving platform to increase diffusion of nutrients provided no additional advantage. Following colonisation by the tumour cells, bone discs pre-seeded with MDA-MB-231 cells were implanted subcutaneously into NOD SCID mice, and tumour growth monitored using in vivo imaging for up to 6 weeks. Tumour growth progressed in human bone discs in 80 % of the animals mimicking the later stages of human bone metastasis. Immunohistochemical and PCR analysis revealed that growing MDA-MB-231 cells in human bone resulted in these cells acquiring a molecular phenotype previously associated with breast cancer bone metastases. MDA-MB-231 cells grown in human bone discs showed increased expression of IL-1B, HRAS and MMP9 and decreased expression of S100A4, whereas, DKK2 and FN1 were unaltered compared with the same cells grown in mammary fat pads of mice not implanted with human bone discs.

Expression and function of enamel-related gene products in calvarial development.

Enamel-related gene products (ERPs) are detected in non-enamel tissues such as bone. We hypothesized that, if functional, ERP expression corresponds with distinct events during osteoblast differentiation and affects bone development and mineralization. In mouse calvariae and MC3T3 cells, expression profiles of enamel-related gene products (ERPs) correlated with key events in post-natal calvarial development and MC3T3 cell mineralization. Developing skulls from both Amel- and Ambn-deficient animals were approximately 15% shorter when compared with those of wild-type controls, and their sutures remained patent for a longer period of time. Analysis of Amel- and Ambn-deficient calvariae and calvarial osteoblast cultures revealed a dramatic reduction in mineralized nodules, a significant reduction in Runx2, Sp7, Ibsp, and Msx2 expression, and a reduction in Alx4 in Amel-deficient calvariae vs. an increase in Alx4 in Ambn-deficient calvariae. Analysis of these data indicates that ERP expression follows defined developmental profiles and affects osteoblast differentiation, mineralization, and calvarial bone development. We propose that, in parallel to their role in the developing enamel matrix, ERPs have retained an evolutionary conserved function related to the biomineralization of bones.

Probiotic viability and physico-chemical and sensory properties of plain and stirred fruit yogurts made from goat's milk.

Probiotic plain and stirred fruit yogurts were made from goat's milk using bacterial cultures comprising, Lactobacillus acidophilus LA-5, Bifidobacterium animalis subsp. lactis BB-12 and Propionibacterium jensenii 702. The products were stored at 4°C for 4weeks, during which time the viability of the yogurt starter culture and probiotic bacteria was analysed weekly. P. jensenii 702 demonstrated the highest viability (10(8)cfu/g) in all types of yogurt throughout the storage period, while the viability of the bifidobacteria (∼10(7)cfu/g) also remained above the minimum therapeutic level. The viability of L. acidophilus LA-5 fell below 10(6)cfu/g in yogurts, however, the addition of fruit juice appeared to support the viability of lactobacilli, with higher microorganism numbers observed in fruit yogurts than in plain yogurt throughout the shelf life. Addition of fruit juice significantly increased the syneresis, and decreased viscosity and water holding capacity of yogurts (p<0.05), and also enhanced their sensory acceptability.

The Colour Test for drug susceptibility testing of Mycobacterium tuberculosis strains.

SETTING: Tartu, Estonia. OBJECTIVE: To assess the performance and feasibility of the introduction of the thin-layer agar MDR/XDR-TB Colour Test (Colour Test) as a non-commercial method of drug susceptibility testing (DST). DESIGN: The Colour Test combines the thin-layer agar technique with a simple colour-coded quadrant format, selective medium to reduce contamination and colorimetric indication of bacterial growth to simplify interpretation. DST patterns for isoniazid (INH), rifampicin (RMP) and ciprofloxacin (CFX) were determined using the Colour Test for 201 archived Mycobacterium tuberculosis isolates. Susceptibilities were compared to blinded DST results obtained routinely using the BACTEC™ Mycobacteria Growth Indicator Tube™ (MGIT) 960 to assess performance characteristics. RESULTS: In all, 98% of the isolates produced interpretable results. The average time to positivity was 13 days, and all results were interpretable. The Colour Test detected drug resistance with 98% sensitivity for INH, RMP and CFX and 99% for multidrug-resistant tuberculosis. Specificities were respectively 100% (95%CI 82-100), 88% (95%CI 69-97) and 91% (95%CI 83-96) and 90% (95%CI 74-98). Agreement between the Colour Test and BACTEC MGIT 960 were respectively 98%, 96%, 94% and 97%. CONCLUSION: The Colour Test could be an economical, accurate and simple technique for testing tuberculosis strains for drug resistance. As it requires little specialist equipment, it may be particularly useful in resource-constrained settings with growing drug resistance rates.