Diatomaceous earth increases the efficacy of Beauveria bassiana against Tribolium castaneum larvae and increases conidia attachment

This research tested the suppressive ability of Beauveria bassiana (Balsamo) Vuillemin alone and in combination with diatomaceous earth against the red flour beetle, Tribolium castaneum (Herbst). Adults did not show a dose response to B. bassiana, and the addition of diatomaceous earth (DE) did not result in a significant increase in mortality. Against larvae, however, DE at 190 mg/kg grain enhanced the efficacy of B. bassiana at all concentrations ranging from 33 to 2,700 mg of conidia per kilogram of grain. The presence of DE resulted in 17- and 16-fold decreases in the median lethal concentration of B. bassiana at 56 and 75% RH, respectively. No significant differences in larval mortality in response to B. bassiana and diatomaceous earth alone or in combination were found between 56 and 75% RH. Conidial attachment to larvae was significantly greater with 190 mg/kg DE than without it. The partial analysis of lipids taken up by DE from the larvae revealed the removal of phospholipids and long-chain fatty acids. These results support the hypothesis that diatomaceous earth enhances the efficacy of B. bassiana against larval T. castaneum, at least in part by damaging the insect cuticle, thus increasing conidial attachment and making nutrients more available to conidia for their germination.     

Topical glucocorticoid therapy directly induces up-regulation of functional CXCR4 on primed T lymphocytes in the aqueous humor of patients with uveitis

Overexpression of the constitutive chemokine receptor CXCR4 has been shown to contribute to the accumulation of leukocytes at sites of chronic inflammation. Glucocorticoids are widely used to treat inflammatory disorders such as uveitis to considerable effect, yet paradoxically have been reported to increase CXCR4 expression in vitro. We show here that ocular lymphocytes isolated from patients with uveitis who had been treated with topical glucocorticoids expressed highly elevated levels of CXCR4. The up-regulation of CXCR4 could be reproduced in vitro by culture of CD4(+) T cells with aqueous humor (AqH), indicating a role for the ocular microenvironment rather than preferential recruitment of CXCR4(+) cells. Untreated uveitis and noninflammatory AqH up-regulated CXCR4 to a limited extent; this was dependent on TGF-beta2. However, the highest levels of CXCR4 both in vivo and in vitro were found in the glucocorticoid-treated patients. Glucocorticoids appeared to be directly responsible for the induction of CXCR4 in treated patients, as the glucocorticoid receptor antagonist RU38486 inhibited the in vitro up-regulation by AqH from these patients. Dexamethasone selectively up-regulated CXCR4 in vitro, but not any of a wide range of other chemokine receptors. CXCL12, the ligand for CXCR4, was present in AqH under noninflammatory conditions, but the levels were low in untreated uveitis and undetectable in treated uveitis AqH. The importance of these results for the treatment of HIV patients with glucocorticoids is discussed as well as a role for glucocorticoid-induced CXCR4 up-regulation and CXCL12 down-regulation in controlling the migration of lymphocyte populations, resulting in resolution of inflammation.     

Human papillomavirus DNA and liquid-based cervical cytology cotesting in screening and follow-up patient groups

OBJECTIVE: To compare the use of human papillomavirus (HPV) DNA and cervical cytology cotesting in screening and follow-up of patients with previous cervical abnormalities and to assess the significance of a positive HPV DNA test result in re-screening of cytologically normal cases. STUDY DESIGN: Cellular samples collected in liquid-based fixative were used for both cervical cytology and HPV DNA testing. The cervical cytology slides were manually screened by cytotechnologists followed by rapid re-screening by pathologists. The HPV DNA tests were performed using hybrid capture test kits. Statistical analyses of cervical cytology results and HPV DNA tests for high- and low-risk HPV from both patient groups were carried out. RESULTS: The prevalence of HPV DNA-positive cases was higher in younger patients. There was a poor correlation between cervical cytology results and HPV DNA tests for the screening group (kappa = 0.23), but a fair to good correlation was obtained for the follow-up group (kappa = 0.51). The false negative fraction of cytology negative/HPV DNA positive cases (0.1317), as compared with cytology negative/HPV DNA negative cases (0.0056), was statistically significant (p = 0.000001). CONCLUSION: The prevalence of HPV DNA decreased with increasing age in both the screening and follow-up patient groups. Virus clearance was delayed in the follow-up group as compared with the screening group. There was a poor correlation between cervical cytology and HPV DNA tests in the screening group but a fair to good correlation in the follow-up patient group. Cotesting of HPV DNA and cervical cytology increases the sensitivity and decreases the false negative fraction, suggesting that cotesting could be used to increase the interval of screening.     

Survival and behavioral characteristics of amphidromous goby larvae of Sicyopterus japonicus (Tanaka, 1909) during their downstream migration

To understand the ecology and environmental tolerances of newly hatched larvae of the amphidromous fish Sicyopterus japonicus during their downstream migration, the salinity tolerance of eggs, 0-15 day old larvae, and adults, and the temperature tolerance, specific gravity and phototaxis of hatched larvae were examined. Tolerances of adults were measured as survival after a 24 h challenge in freshwater (FW), brackish water (1/3 SW) and seawater (SW). The survival rate of adult S. japonicus was 100% in FW and 1/3 SW, while none survived in SW. Hatching success of eggs (30 eggs each) was significantly higher in FW (mean: 73%) and 1/3 SW (73%) than in SW (19%). Tolerance of newly hatched larvae to salinity and temperature was investigated in different combinations of salinities (FW, 1/3 SW and SW) and temperatures (18, 23 and 28 °C). Larval survival was significantly different in each salinity and temperature. Survival rate was significantly higher in 1/3 SW than in FW and higher in SW than in FW at 23 °C and 28 °C. At the latter part of the experiment, there was no survival in FW and at 28 °C. Survival was higher in lower temperatures, but larval development did not occur in FW. Specific gravity of newly hatched larvae was 1.036 at 28 °C and 1.034 at 23 °C. When exposed to a light source on one side of an aquarium, larval distribution was not affected. Our results indicated larval S. japonicus are more adapted to brackish water and seawater than freshwater, while the adults and eggs are more adapted to freshwater and brackish water than seawater. This is consistent with their amphidromous life history with growth and spawning occurring in freshwater and the larval stage utilizing marine habitats.     

Identification of a novel human granzyme B inhibitor secreted by cultured sertoli cells

Sertoli cells have long since been recognized for their ability to suppress the immune system and protect themselves as well as other cell types from harmful immune reaction. However, the exact mechanism or product produced by Sertoli cells that affords this immunoprotection has never been fully elucidated. We examined the effect of mouse Sertoli cell-conditioned medium on human granzyme B-mediated killing and found that there was an inhibitory effect. We subsequently found that a factor secreted by Sertoli cells inhibited killing through the inhibition of granzyme B enzymatic activity. SDS-PAGE analysis revealed that this factor formed an SDS-insoluble complex with granzyme B. Immunoprecipitation and mass spectroscopic analysis of the complex identified a proteinase inhibitor, serpina3n, as a novel inhibitor of human granzyme B. We cloned serpina3n cDNA, expressed it in Jurkat cells, and confirmed its inhibitory action on granzyme B activity. Our studies have led to the discovery of a new inhibitor of granzyme B and have uncovered a new mechanism used by Sertoli cells for immunoprotection.     

Nutrient stress and performance of invasive Hieracium lepidulum and co-occurring species in New Zealand

Many studies have highlighted the relationship between nutrient fluctuations and enrichment in the process of plant invasion. However, invasion may be also associated with conditions of plant stress, either nutrient depletion or toxicity, in the environment. In this study, we investigate the possible role of nutrient stress in the invasion of Hieracium lepidulum (Stenstroem) Omang, in South Island, New Zealand. We do this by comparing several performance attributes, and their plasticity, for H. lepidulum and a number of co-occurring species, across a series of nutrient depletion and toxicity tests. H. lepidulum had intermediate yields, high root:shoot ratios and high tissue nutrient contents at control nutrient concentrations. H. lepidulum differed in edaphic tolerance from all but Chionochloa flavescens var. brevis, in being insensitive to nutrient dilutions other than nitrogen. The significance of performance in terms of edaphic tolerance and adaptations are discussed. Intermediate yields at control nutrient levels suggest that H. lepidulum should not be competitive compared to high yielding species including Agrostis stolonifera and Poa cita, but should be competitive with lower yielding Coprosma rugosa and C. flavescens var. brevis. Conversely, significant yield decreases under nitrogen limitation stress suggests that H. lepidulum will not likely occupy very nutrient poor sites. H. lepidulum, along with C. flavescens var. brevis, were found to be tolerant of ammonium as an alternative source of nitrogen, while other species were not. These data suggest that H. lepidulum and C. flavescens var. brevis would be relatively tolerant of the stresses associated with acidic soils compared to the other species, but not to stresses associated with absolute shortage of nitrogen. Combined results point to the likely occurrence of H. lepidulum at sites of intermediate fertility. The possible roles of ammonium stress and disturbance reliance in further defining H. lepidulum ecology is discussed.     

Exogenous free ubiquitin enhances lily pollen tube adhesion to an in vitro stylar matrix and may facilitate endocytosis of SCA

Pollen tube adhesion and guidance on extracellular matrices within the pistil are essential processes that convey the pollen tube cell and the sperm cells to the ovule. In this study, we purified an additional molecule from the pistil that enhances pollen tube adhesion when combined with the SCA (stigma/stylar cysteine-rich adhesin)/pectin matrix in our in vitro assay. The enhancer of adhesion was identified as free ubiquitin (Ub). This was confirmed by use of bovine Ub as a substitute for lily (Lilium longiflorum Thunb.) stigma Ub. To study the interaction of SCA and Ub with the lily pollen tube, we labeled both proteins with biotin. We observed uptake of biotin-labeled SCA and Ub into the pollen tube cells in vitro using confocal microscopy. For SCA, a strong signal occurred first at the tip of the pollen tube, suggestive of an endocytosis event, and then progressively throughout the tube cytoplasm. SCA was also localized inside the in vivo pollen tube using immunogold electron microscopy and found to be present in endosomes, multivesicular bodies, and vacuoles, all known to be endocytic compartments. It was also confirmed that SCA is endocytosed in the in vitro adhesion assay. Internalization of SCA was increased in pollen tubes treated with exogenous Ub compared to those without Ub, suggesting that Ub may facilitate SCA endocytosis. These results show that Ub can act as an enhancer of pollen tube adhesion in vitro and that it is taken up into the pollen tube as is SCA. The Ub machinery may play a role in pollen tube adhesion and guidance in lily.     

Comparison of the genome sequence of the poultry pathogen Bordetella avium with those of B. bronchiseptica, B. pertussis, and B. parapertussis reveals extensive diversity in surface structures associated with host interaction

Bordetella avium is a pathogen of poultry and is phylogenetically distinct from Bordetella bronchiseptica, Bordetella pertussis, and Bordetella parapertussis, which are other species in the Bordetella genus that infect mammals. In order to understand the evolutionary relatedness of Bordetella species and further the understanding of pathogenesis, we obtained the complete genome sequence of B. avium strain 197N, a pathogenic strain that has been extensively studied. With 3,732,255 base pairs of DNA and 3,417 predicted coding sequences, it has the smallest genome and gene complement of the sequenced bordetellae. In this study, the presence or absence of previously reported virulence factors from B. avium was confirmed, and the genetic bases for growth characteristics were elucidated. Over 1,100 genes present in B. avium but not in B. bronchiseptica were identified, and most were predicted to encode surface or secreted proteins that are likely to define an organism adapted to the avian rather than the mammalian respiratory tracts. These include genes coding for the synthesis of a polysaccharide capsule, hemagglutinins, a type I secretion system adjacent to two very large genes for secreted proteins, and unique genes for both lipopolysaccharide and fimbrial biogenesis. Three apparently complete prophages are also present. The BvgAS virulence regulatory system appears to have polymorphisms at a poly(C) tract that is involved in phase variation in other bordetellae. A number of putative iron-regulated outer membrane proteins were predicted from the sequence, and this regulation was confirmed experimentally for five of these.     

The association of Shiga-like toxin with detergent-resistant membranes is modulated by glucosylceramide and is an essential requirement in the endoplasmic reticulum for a cytotoxic effect

Receptor-mediated internalization to the endoplasmic reticulum (ER) and subsequent retro-translocation to the cytosol are essential sequential processes required for the productive intoxication of susceptible mammalian cells by Shiga-like toxin-1 (SLTx). Recently, it has been proposed that the observed association of certain ER-directed toxins and viruses with detergent-resistant membranes (DRM) may provide a general mechanism for their retrograde transport to endoplasmic reticulum (ER). Here, we show that DRM recruitment of SLTx bound to its globotriosylceramide (Gb(3)) receptor is mediated by the availability of other glycosphingolipids. Reduction in glucosylceramide (GlcCer) levels led to complete protection against SLTx and a reduced cell surface association of bound toxin with DRM. This reduction still allowed efficient binding and transport of the toxin to the ER. However, toxin sequestration within DRM of the ER was abolished under reduced GlcCer conditions, suggesting that an association of toxin with lipid microdomains or rafts in the ER (where these are defined by detergent insolubility) is essential for a later step leading to or involving retro-translocation of SLTx across the ER membrane. In support of this, we show that a number of ER residents, proteins intimately involved in the process of ER dislocation of misfolded proteins, are present in DRM.