Seroprevalence of Mycoplasma bovis in bulk milk samples in Irish dairy herds and risk factors associated with herd seropositive status.

Mycoplasma bovis is a serious disease of cattle worldwide; mastitis, pneumonia, and arthritis are particularly important clinical presentations in dairy herds. Mycoplasma bovis was first identified in Ireland in 1994, and the reporting of Mycoplasma-associated disease has substantially increased over the last 5 years. Despite the presumed endemic nature of M. bovis in Ireland, there is a paucity of data on the prevalence of infection, and the effect of this disease on the dairy industry. The aim of this observational study was to estimate apparent herd prevalence for M. bovis in Irish dairy herds using routinely collected bulk milk surveillance samples and to assess risk factors for herd seropositivity. In autumn 2018, 1,500 herds out of the 16,858 herds that submitted bulk tank milk (BTM) samples to the Department of Agriculture testing laboratory for routine surveillance were randomly selected for further testing. A final data set of 1,313 sampled herds with a BTM ELISA result were used for the analysis. Testing was conducted using an indirect ELISA kit (ID Screen Mycoplasma bovis). Herd-level risk factors were used as explanatory variables to determine potential risk factors associated with positive herd status (reflecting past or current exposure to M. bovis). A total of 588 of the 1,313 BTM samples were positive to M. bovis, providing an apparent herd prevalence of 0.45 (95% CI: 0.42, 0.47) in Irish dairy herds in autumn 2018. Multivariable analysis was conducted using logistic regression. The final model identified herd size, the number of neighboring farms, in-degree and county as statistically significant risk factors for herd BTM seropositivity to M. bovis. The results suggest a high apparent herd prevalence of seropositivity to M. bovis, and evidence that M. bovis infection is now endemic in the Irish dairy sector. In addition, risk factors identified are closely aligned to what we would expect of an infectious disease. Awareness raising and education about this important disease is warranted given the widespread nature of exposure and likely infection in Irish herds. Further work on the validation of diagnostic tests for herd-level diagnosis should be undertaken as a matter of priority.

Significant re-emergence and recirculation of Schmallenberg virus in previously exposed dairy herds in Ireland in 2016.

Schmallenberg virus (SBV) circulation was investigated in 25 previously exposed dairy herds in Ireland in 2016. A population of 1,550 spring-2014-born animals, which had been monitored for SBV infection in 2014 and 2015 as part of a previous SBV surveillance study, were resampled for evidence of SBV infection during 2016. A total of 366 blood samples were collected in the 25 study herds (15 samples per herd) between 3 March 2017 and 10 March 2017 (before the 2017 vector-active season) and analysed for SBV antibodies using a competitive ELISA kit (IDVet). A total of 256 animals tested seropositive, an AP of 69.9% (95% CI: 65.1-74.4) and TP of 77.7% (95% CI: 72.3%-82.8%) when correcting for imperfect test characteristics. These results demonstrate that a new epidemic of SBV circulation occurred in these previously exposed herds in Ireland in 2016.

Offsetting the impacts of mining to achieve no net loss of native vegetation.

Offsets are a novel conservation tool, yet using them to achieve no net loss of biodiversity is challenging. This is especially true when using conservation offsets (i.e., protected areas) because achieving no net loss requires avoiding equivalent loss. Our objective was to determine if offsetting the impacts of mining achieves no net loss of native vegetation in Brazil's largest iron mining region. We used a land-use change model to simulate deforestation by mining to 2020; developed a model to allocate conservation offsets to the landscape under 3 scenarios (baseline, no new offsets; current practice, like-for-like [by vegetation type] conservation offsetting near the impact site; and threat scenario, like-for-like conservation offsetting of highly threatened vegetation); and simulated nonmining deforestation to 2020 for each scenario to quantify avoided deforestation achieved with offsets. Mines cleared 3570 ha of native vegetation by 2020. Under a 1:4 offset ratio, mining companies would be required to conserve >14,200 ha of native vegetation, doubling the current extent of protected areas in the region. Allocating offsets under current practice avoided deforestation equivalent to 3% of that caused by mining, whereas allocating under the threat scenario avoided 9%. Current practice failed to achieve no net loss because offsets did not conserve threatened vegetation. Explicit allocation of offsets to threatened vegetation also failed because the most threatened vegetation was widely dispersed across the landscape, making conservation logistically difficult. To achieve no net loss with conservation offsets requires information on regional deforestation trajectories and the distribution of threatened vegetation. However, in some regions achieving no net loss through conservation may be impossible. In these cases, other offsetting activities, such as revegetation, will be required.

Sensitivity of host-seeking nymphal lone star ticks (Acari: Ixodidae) to immersion in heated water.

Host-seeking nymphal Amblyomma americanum (L.) (Acari: Ixodidae) were placed into heated water, and their survival or their torpidity was recorded as a function of exposure time. Exposures were determined that either kill the nymphs or affect their mobility. All nymphs died when exposed for a minute or more to a temperature > 51 degrees C. Nearly all nymphs remained motionless for a period of time when exposed for 3 min to a temperature > 44 degrees C.

Some investigations into non-passive listening.

Our knowledge of the function of the auditory nervous system is based upon a wealth of data obtained, for the most part, in anaesthetised animals. More recently, it has been generally acknowledged that factors such as attention profoundly modulate the activity of sensory systems and this can take place at many levels of processing. Imaging studies, in particular, have revealed the greater activation of auditory areas and areas outside of sensory processing areas when attending to a stimulus. We present here a brief review of the consequences of such non-passive listening and go on to describe some of the experiments we are conducting to investigate them. In imaging studies, using fMRI, we can demonstrate the activation of attention networks that are non-specific to the sensory modality as well as greater and different activation of the areas of the supra-temporal plane that includes primary and secondary auditory areas. The profuse descending connections of the auditory system seem likely to be part of the mechanisms subserving attention to sound. These are generally thought to be largely inactivated by anaesthesia. However, we have been able to demonstrate that even in an anaesthetised preparation, removing the descending control from the cortex leads to quite profound changes in the temporal patterns of activation by sounds in thalamus and inferior colliculus. Some of these effects seem to be specific to the ear of stimulation and affect interaural processing. To bridge these observations we are developing an awake behaving preparation involving freely moving animals in which it will be possible to investigate the effects of consciousness (by contrasting awake and anaesthetized), passive and active listening.

A study of dry cow therapy and effects on SCC in 10 Irish dairy herds.

Somatic cell count (SCC) data for 480 cows in 10 Irish dairy herds from January 2001 until June 2002 were analysed. Herds were selected on the basis of a recent or ongoing history of clinical or subclinical mastitis. An individual cow SCC of 200 000 cells per ml was used as the threshold for elevation of SCC. The duration of elevated SCC prior to drying-off and the magnitude of the elevation in SCC were found to have an impact on the response to dry cow therapy (DCT). A trend also emerged indicating that increasing parity had a negative influence on the response to DCT.

Reflexive shifts of covert attention operate in an egocentric coordinate frame.

Covert shifts of attention have been shown to improve detection and discrimination thresholds for a range of visual stimuli. Although there is some evidence to suggest that the allocation of attention to a particular region of interest occurs in a retinotopic frame of reference, the importance of an allocentric, or object-based, framework has gained widespread empirical support. The current experiment investigates the nature of the spatial representation in which covert shifts of attention occur in response to a reflexive prime. Primes and targets were presented in four conditions designed to vary systematically the validity of the spatial relationship between the prime and target in egocentric or allocentric coordinate frameworks. A significant advantage, in terms of reaction time and correct identification, was found for targets located in positions previously primed in an egocentric (but not allocentric) framework whereas there was no advantage for locations primed in an allocentric (but not egocentric) framework. These results suggest that the allocation of covert spatial attention within an egocentric framework may be more important than previously thought.

Analysis of intensive care populations to select possible candidates for high dependency care.

OBJECTIVES: To identify the proportion, and range across intensive care units, of intensive care patients who might potentially be managed on a high dependency unit (HDU) using three different classification systems. METHODS: 8095 adult patients admitted to 15 intensive care units in the south of England between 1 April 1993 and 31 December 1994 were studied. Patients were identified as potential HDU admissions if their APACHE III derived risk of hospital mortality was < or =10%, if they were categorised as a low risk monitor (LRM) patient using the Wagner risk stratification method, or if they did not require advanced respiratory support (ARS). RESULTS: 4146 patients (51.2%) had an APACHE III derived risk of hospital death of < or =10%, 1687 (20.8%) were classified as LRM, and 3860 (47.7%) did not receive ARS. The values for each intensive care unit ranged from 32.8-63.3% (APACHE III group), 7.2-29.9% (LRM group), and 14.4-68.2% (ARS group). No matter which of the three methods was used, there were significant differences between the 15 units (p<0.0001) with regard to the number of potential HDU patients identified within the scored population. CONCLUSIONS: The percentage of intensive care patients who might be more appropriately managed in a HDU varies considerably between hospitals, and depends upon both local circumstances and the method used to define a high dependency patient. However, whichever method is used, it appears that significant numbers of patients of low dependency status currently fill intensive care beds in the units studied. If these analyses are correct, the perceived national shortage in intensive care beds might be improved by the development of HDUs.

Sensitization of multidrug-resistant human leukemia cells with MDR1-targeted antisense and inhibition of drug-mediated MDR1 induction.

Increased expression of MDR1 is strongly implicated in the appearance of chemotherapeutic drug resistance in cancer, especially hematological malignancies. We therefore examined the potential of antisense oligonucleotides to inhibit MDR1 and restore sensitivity to drug-resistant human lymphoblastic cells (CCRF-CEM). Treatment with two different phosphorothioate-modified antisense sequences as well as a DNA-RNA hybrid sequence resulted in a 30 to 45% decrease in MDR1 expression as determined by staining with the monoclonal antibody MRK16 followed by flowcytometry (FCM) analysis. Further, inhibition of MDR1 expression persisted for 3 days after removal of oligonucleotides. Increased accumulation of rhodamine 123 and nearly a three-fold sensitization of cells to vincristine paralleled the reduction in staining with MRK16. Reversed or scrambled control sequences had no effect in any of the assays. During the course of these studies, we observed a 25 to 75% increase in MRK16 staining of cells treated with the chemotherapeutic agents daunorubicin and vincristine as well as by the resistance reversal agents verapamil and cyclosporin. Treatment of cells with antisense oligonucleotides prior to exposure to daunorubicin or cyclosporin reduced the increase in MRK16 staining. These results indicate that antisense targeted to MDR1 can sensitize drug-resistant leukemia cells and suggest that antisense treatment may prevent the emergence of MDR1-mediated drug resistance.

Hsp90-mediated folding of the lymphoid cell kinase p56lck.

Several lines of evidence suggest that members of the 90-kDa family of heat shock proteins (hsp90) may support the folding of various homologues of the src kinase family. In this work, we utilized pulse-chase analyses in rabbit reticulocyte lysate to demonstrate that hsp90-bound intermediates existed for the majority of newly synthesized p56lck molecules. The hsp90-binding drug geldanamycin disrupted the association of p56lck with hsp90, prevented the kinase from demonstrating a protease-resistant conformation, and caused decreases in kinase specific activity. Requirements for geldanamycin-inhibitable hsp90 function and physical interactions between hsp90 and p56lck persisted during chase periods. Consistent with the effects observed in rabbit reticulocyte lysate, application of geldanamycin to fibroblasts caused specific reversion of lck-mediated transformation concomitant with loss of p56lck activity and protein. However, geldanamycin had no direct effect on purified p56lck. Also consistent with functional linkages between hsp90 and p56lck, physical interactions between these proteins were detected in cytoplasmic, but not membrane, fractions of LSTRA cells. Although hsp90 functions in both the initial de novo folding and the reiterative support of p56lck structure in rabbit reticulocyte lysate, the specific occurrence of complexes between hsp90 and p56lck in the cytoplasm of T cells suggests that hsp90 primarily folds nascent molecules of p56lck in vivo.

The outcome of poliovirus infections in K562 cells is cytolytic rather than persistent after hemin-induced differentiation.

K562-Mu erythroleukemia cells readily establish a long-term persistent poliovirus infection characterized by continuous virus production in the absence of complete p220 cleavage and host translation shutoff (R. E. Lloyd and M. Bovee, Virology 194:200-209, 1993). The mechanism of resistance appears to be modulated at the intracellular level and to be related to decreased virus-mediated cytopathic effects (P. A. Benton, J. W. Murphy, and R. E. Lloyd Virology 213:7-18, 1995). It is well documented that hemin induces the differentiation of K562 cells and alters the expression of several host proteins. We report here that growth of K562 cells in hemin prior to poliovirus infection results in a dose-dependent increase in virus-induced cell lysis and thereby alters the normally persistent outcome of infection to a more lytic phenotype. K562 cells infected after hemin treatment displayed increased host translation shutoff, p220 cleavage, viral protein synthesis, and viral RNA accumulation compared with nontreated cells. Since hemin treatment of K562 cells also induced the increased expression of several heat shock proteins (Hsp70, Hsc70, Hsp90, and cohort p60), we tested the hypothesis that their increased expression may play a role in altering poliovirus infection in hemin-treated K562 cells. However, neither heat stress nor oxidative stress, inducers of heat shock protein synthesis, altered the outcome (of virus infections. In addition, we report the novel finding that subunits of two translation initiation factors, p220 (eIF-4G) and eIF-2alpha, are cleaved as a result of hemin treatment of K562 cells. It is proposed that hemin alters the expression of specific host proteins in K562 cells, probably other than heat shock proteins, which changes the initial response to poliovirus infections from persistent to lytic.

Arrested rearrangement of TCR V beta genes in thymocytes from children with X-linked severe combined immunodeficiency disease.

Human X-linked severe combined immunodeficiency disease (SCID) is an immunodeficiency disorder in which T cell development is arrested in the thymic cortex. B lymphocytes in children with X-linked SCID seem to differentiate normally. X-linked SCID is associated with a mutation in the gene that encodes the IL-2R gamma-chain. Because TCR-beta gene recombination is a pivotal initial event in T lymphocyte ontogeny within the thymus, we hypothesized that a failure to express normal IL-2R gamma could lead to impaired TCR-beta gene recombination in early thymic development. PCR was used to determine the status of TCR-beta gene-segment rearrangements in thymic DNA that had been obtained from children with X-linked SCID. The initial step in TCR-beta gene rearrangement, that of D beta to J beta recombination, was readily detected in all thymus samples from children with X-linked SCID; in contrast, V beta to DJ beta gene rearrangements were undetectable in the same samples. Both D beta to J beta and V beta to DJ beta TCR genes were rearranged in the thymic tissues obtained from immunologically normal children. We conclude that TCR beta-chain gene rearrangement is arrested in children with X-linked SCID. Our results suggest a causative relationship between the failure of TCR beta-chain gene rearrangements to proceed beyond DJ beta rearrangements and the production of a nonfunctional IL-2R gamma-chain.

Persistence of multiple maternal genotypes of human immunodeficiency virus type I in infants infected by vertical transmission.

The extent of nucleotide variation within the HIV-1 env hypervariable domains serves as a marker of virus genotypes within infected individuals and as a means to track transmission of the virus between individuals. We analyzed env V1 and V2 sequences in longitudinal samples from two HIV-1-infected mothers, each with three children infected by maternal transmission of the virus. Sequences in samples that were obtained from two infants at 2 d and 4 wk after birth displayed more variation in V1 and V2 than maternal samples obtained at the same times. Multiple HIV-1 genotypes were identified in each mother. In each family, multiple maternal HIV-1 genotypes were transmitted to the infants. Specific amino acid residues in the hypervariable domains were conserved within sequences from each family producing a family-specific amino acid signature pattern in V1 and V2. Viruses that were highly related to maternal viruses in signature pattern persisted for as long as 4 yr in the older children. Results support a model of transmission involving multiple HIV-1 genotypes with development of genetic variation from differential outgrowth and accumulation of genetic changes within each individual.

Independent variation and positive selection in env V1 and V2 domains within maternal-infant strains of human immunodeficiency virus type 1 in vivo.

Multiple targets for immune recognition and cellular tropism are localized to the V1 and V2 hypervariable regions in the amino portion of human immunodeficiency virus type 1 (HIV-1) gp120env. We have assessed genetic diversity in env V1 and V2 hypervariable domains in vivo within epidemiologically related strains of HIV-1. Our strategy was to analyze longitudinal samples from two seropositive mothers and multiple children infected by perinatal transmission. Although the V1 and V2 domains are closely linked in the HIV-1 genome, nucleotide sequences in V1 and in V2 evolved independently in maternal-infant viruses in vivo. A high proportion of the nucleotide substitutions would introduce amino acid diversity in V1 and in V2. A significant excess of nonsynonymous over synonymous substitutions was identified in HIV-1 env V1 and V2 peptides in the mothers and in two older children but was not generally apparent in HIV-1 sequences in infants. An excess of nonsynonymous over synonymous substitutions indicated that there is positive selection for independent genetic variation in the V1 and V2 domains in vivo. It is likely that there are host responses to complex determinants in the V1 or V2 hypervariable domain of HIV-1 gp120.

Ocular manifestations of the lacrimo-auriculo-dento-digital syndrome.

We studied a mother and daughter with an extremely rare constellation of signs and symptoms. One or both had absent lacrimal puncta, nasolacrimal duct obstruction, chronic dacryocystitis, dry eyes, and epiphora. Systemic findings included salivary gland hyposecretion, dental hypoplasia and dysplasia, cup-shaped ears with hearing loss, and digital anomalies. These findings are consistent with those of the lacrimo-auriculo-dento-digital syndrome, a genetic disorder. Our study supports the autosomal dominant inheritance of this syndrome, delineates the ophthalmic manifestations, and provides evidence that renal anomalies are part of the disorder.

The temporal association between gamma delta T cells and the natural history of insulin-dependent diabetes.

We previously reported a significant increase in percentages of peripheral blood gamma delta+ T cells in islet cell antibody (ICA) positive relatives of patients with insulin-dependent diabetes (IDD). In the present study, we further characterized this T cell abnormality in a larger group of ICA+ subjects and report that (1) Percentages of gamma delta+ T lymphocytes were significantly increased only in subjects with high ICA titers (> or = 20 JDF units) (P = 0.005) and resulted from an increase in absolute numbers of gamma delta+ T lymphocytes. (2) In these subjects, the increase in gamma delta+ T lymphocytes was associated with an increase in the V gamma 9 V delta 2 subpopulation (r = 0.99). (3) In these same subjects, high percentages of gamma delta+ T lymphocytes were associated with normal beta cell function while low percentages were associated with diminished insulin response. Using 65 microU/ml as the threshold of abnormal intravenous glucose tolerance test (IVGTT) response, percentages of gamma delta+ T lymphocytes could significantly predict IVGTT status in these subjects (P < 0.01). A longitudinal follow-up further suggested that the development of an abnormal IVGTT response and progression to diabetes was associated with a decrease in percentages of gamma delta+ T lymphocytes while patients whose gamma delta+ T cell percentages remained high retained normal beta cell function. Our data therefore suggest that gamma delta+ T lymphocytes and more specifically V gamma 9 V delta 2 T cells are implicated in the autoimmune process leading to diabetes and may have a regulatory role. The monitoring of their percentages in the blood of patients at risk for diabetes may be useful as an additional predictor of diabetes development.

Corticosteroids improve survival of children with AIDS and Pneumocystis carinii pneumonia.

OBJECTIVE: To determine if corticosteroids administered in addition to antimicrobials improve survival in children with acquired immunodeficiency syndrome and severe Pneumocystis carinii pneumonia (PCP). DESIGN: Before-after, nonrandomized, case-comparison study. SETTING: Pediatric intensive care unit of a tertiary care teaching hospital in Florida. PARTICIPANTS: Eleven children infected with the human immunodeficiency virus (HIV) with confirmed PCP. SELECTION PROCEDURE: Infants with HIV infection and acute respiratory failure due to PCP were studied sequentially. INTERVENTION: The first seven infants were treated with antimicrobials alone while the next four received a 2-week course of methylprednisolone sodium succinate in addition to antimicrobials. MEASUREMENTS AND RESULTS: The two groups were similar with respect to age, route of HIV infection, stage of HIV disease, CD4 T-cell count, antiretroviral therapy, and respiratory parameters at intubation. All children treated with antimicrobials alone died while receiving mechanical ventilation. Survival of the children who received corticosteroid therapy was significantly improved (P < .05), and all were weaned from the ventilator. CONCLUSIONS: Corticosteroids administered in addition to antimicrobials and supportive care improve the short-term survival of HIV-infected children who have acute respiratory failure due to PCP.

Human anti-pneumococcal polysaccharide antibodies are secreted by the CD5- B cell lineage.

To determine whether human antibody responses to T cell-independent pneumococcal polysaccharide antigens are derived from CD5+ or CD5- B cells, we utilized an ELISPOT assay to detect individual anti-polysaccharide antibody-secreting cells. Human anti-type IV pneumococcal polysaccharide antibody-secreting cells were found in the CD5- B cell subpopulation. An EBV transformed anti-pneumococcal antibody-secreting B cell line was also CD5-. The ontogeny of CD5 expressing B cells correlated with the age at which polysaccharide responsiveness is acquired (generally around age 2 years in humans). The CD5- B cell subset represents only 25-30% of the B cells in young children, but this fraction increases throughout childhood to a plateau of 70-80% of the B cells in adults. These results support the hypothesis that the developmental change in responsiveness to T cell-independent polysaccharide antigens in humans is associated with maturation of the CD5- B cell subset.