Pore-Forming Toxins Induce Macrophage Necroptosis during Acute Bacterial Pneumonia

Necroptosis is a highly pro-inflammatory mode of cell death regulated by RIP (or RIPK)1 and RIP3 kinases and mediated by the effector MLKL. We report that diverse bacterial pathogens that produce a pore-forming toxin (PFT) induce necroptosis of macrophages and this can be blocked for protection against Serratia marcescens hemorrhagic pneumonia. Following challenge with S. marcescens, Staphylococcus aureus, Streptococcus pneumoniae, Listeria monocytogenes, uropathogenic Escherichia coli (UPEC), and purified recombinant pneumolysin, macrophages pretreated with inhibitors of RIP1, RIP3, and MLKL were protected against death. Alveolar macrophages in MLKL KO mice were also protected during S. marcescens pneumonia. Inhibition of caspases had no impact on macrophage death and caspase-1 and -3/7 were determined to be inactive following challenge despite the detection of IL-1β in supernatants. Bone marrow-derived macrophages from RIP3 KO, but not caspase-1/11 KO or caspase-3 KO mice, were resistant to PFT-induced death. We explored the mechanisms for PFT-induced necroptosis and determined that loss of ion homeostasis at the plasma membrane, mitochondrial damage, ATP depletion, and the generation of reactive oxygen species were together responsible. Treatment of mice with necrostatin-5, an inhibitor of RIP1; GW806742X, an inhibitor of MLKL; and necrostatin-5 along with co-enzyme Q₁₀ (N5/C₁₀)_, which enhances ATP production; reduced the severity of S. marcescens pneumonia in a mouse intratracheal challenge model. N5/C₁₀ protected alveolar macrophages, reduced bacterial burden, and lessened hemorrhage in the lungs. We conclude that necroptosis is the major cell death pathway evoked by PFTs in macrophages and the necroptosis pathway can be targeted for disease intervention.     

Chemical characterization and quantification of siderophores produced by marine and terrestrial aspergilli

Ten aspergilli (five each from marine and terrestrial habitats) were screened for siderophore production. All test isolates produced siderophores as indicated by a positive reaction in the FeCl(3) test, chrome azurol sulphonate assay, and chrome azurol sulphonate agar plate test. Further, the test isolates were compared for their siderophore production potential and chemical characteristics. Examination of the chemical nature of the siderophores revealed that all test isolates produced hydroxamate siderophores that were trihydroxamate hexadentates. Wide-spread occurrence of siderophores in marine isolates indicate their functional role in maintaining overall productivity of coastal waters. Among all test aspergilli, marine Aspergillus versicolor was found to be the largest siderophore producer (182.5 microg/mL desferrioxamine mesylate equivalent), least siderophore production was recorded in a marine strain of Aspergillus niger (3.5 microg/mL desferrioxamine mesylate equivalent).     

Response of Faidherbia albida (Del.) A. Chev., Acacia nigrescens Oliver. and Acacia nilotica (L.) Willd ex Del. seedlings to simulated cotyledon and shoot herbivory in a semi‐arid savanna in Zimbabwe

Woody plant seedling establishment is constrained by herbivory in many semi‐arid savannas. We clipped shoots and cotyledons of three woody species 5‐day (=‘early’) or 28‐day (= ‘late’) post‐emergence to simulate herbivory. Seedlings had shoot apex, one or two cotyledon(s) removed, or were retained intact. Survival rates were ≥80%, ≥40% and ≥20% for Acacia nilotica, Acacia nigrescens and Faidherbia albida respectively. F. albida mobilized stored cotyledon reserves faster and consequently shed the cotyledons earlier than the two Acacia species. Cotyledons were shed off as late as 70 days post‐emergence with 5‐day shedding earlier than 28‐day and cotyledon life‐span decreasing with intensity of defoliation. Shoot apex removal 28‐day resulted in higher compensatory growth than 5‐day in all three species. Cotyledon removal had no effect on shoot length, while shoot apex removal reduced shoot length. In F. albida root growth was stimulated by shoot apex removal. We conclude that potential tolerance to herbivory in terms of seedling survival was of the order A. nilotica > A. nigrescens > F. albida, timing of shoot apex and cotyledon removal influenced seedling growth in terms of biomass and that shoot apex removal stimulated compensatory growth which is critical to seedling survival.     

Pesticide exposure: the hormonal function of the female reproductive system disrupted?

Some pesticides may interfere with the female hormonal function, which may lead to negative effects on the reproductive system through disruption of the hormonal balance necessary for proper functioning. Previous studies primarily focused on interference with the estrogen and/or androgen receptor, but the hormonal function may be disrupted in many more ways through pesticide exposure. The aim of this review is to give an overview of the various ways in which pesticides may disrupt the hormonal function of the female reproductive system and in particular the ovarian cycle. Disruption can occur in all stages of hormonal regulation: 1. hormone synthesis; 2. hormone release and storage; 3. hormone transport and clearance; 4. hormone receptor recognition and binding; 5. hormone postreceptor activation; 6. the thyroid function; and 7. the central nervous system. These mechanisms are described for effects of pesticide exposure in vitro and on experimental animals in vivo. For the latter, potential effects of endocrine disrupting pesticides on the female reproductive system, i.e. modulation of hormone concentrations, ovarian cycle irregularities, and impaired fertility, are also reviewed. In epidemiological studies, exposure to pesticides has been associated with menstrual cycle disturbances, reduced fertility, prolonged time-to-pregnancy, spontaneous abortion, stillbirths, and developmental defects, which may or may not be due to disruption of the female hormonal function. Because pesticides comprise a large number of distinct substances with dissimilar structures and diverse toxicity, it is most likely that several of the above-mentioned mechanisms are involved in the pathophysiological pathways explaining the role of pesticide exposure in ovarian cycle disturbances, ultimately leading to fertility problems and other reproductive effects. In future research, information on the ways in which pesticides may disrupt the hormonal function as described in this review, can be used to generate specific hypotheses for studies on the effects of pesticides on the ovarian cycle, both in toxicological and epidemiological settings.     

Ferric reductase, superoxide dismutase and alkaline phosphatase activities in siderophore producing fungi

Enzymes associated with release of iron from internalized ferrated siderophore (ferrisiderophore reductase), with damage to the cell at high iron concentration (superoxide dismutase) and siderophore synthesis (alkaline phosphatase), were examined in 3 test fungi viz., Aspergillus sp. ABp4, Aureobasidium pullulans and Rhizopus sp. Extracellular ferrisiderophore reductase activity was present in all the three fungi, but Aureobasidium pullulans, that showed the highest activity (84.3 microM min(-1)), was the only one to produce intra-cellular ferric reductase (147.9 microM min(-1)). Superoxide dismutase was produced by Aureobasidium pullulans and Rhizopus sp., but not by Aspergillus sp. ABp4, that showed intra-cellular enzyme activity in case of ferric reductase and alkaline phosphatase. Maximum SOD activity was seen in Aureobasidium pullulans both extra-cellularly (93.83 ng ml(-1)) and intra-cellularly (57.14 ng ml(-1)). All the test fungi examined, produced intra-cellular alkaline phosphatase. There was no extracellular alkaline phosphatase. Among the three fungi, Aureobasidium pullulans showed highest alkaline phosphatase activity (129.9 microM min(-1)) and Aspergillus sp. ABp4 the least (76.4 microM min(-1)).