Maduramicin Rapidly Eliminates Malaria Parasites and Potentiates the Gametocytocidal Activity of the Pyrazoleamide PA21A050

New strategies targeting Plasmodium falciparum gametocytes, the sexual-stage parasites that are responsible for malaria transmission, are needed to eradicate this disease. Most commonly used antimalarials are ineffective against P. falciparum gametocytes, allowing patients to continue to be infectious for over a week after asexual parasite clearance. A recent screen for gametocytocidal compounds demonstrated that the carboxylic polyether ionophore maduramicin is active at low nanomolar concentrations against P. falciparum sexual stages. In this study, we showed that maduramicin has an EC₅₀ (effective concentration that inhibits the signal by 50%) of 14.8 nM against late-stage gametocytes and significantly blocks in vivo transmission in a mouse model of malaria transmission. In contrast to other reported gametocytocidal agents, maduramicin acts rapidly in vitro, eliminating gametocytes and asexual schizonts in less than 12 h without affecting uninfected red blood cells (RBCs). Ring stage parasites are cleared by 24 h. Within an hour of drug treatment, 40% of the normally crescent-shaped gametocytes round up and become spherical. The number of round gametocytes increases to >60% by 2 h, even before a change in membrane potential as monitored by MitoProbe DiIC1 (5) is detectable. Maduramicin is not preferentially taken up by gametocyte-infected RBCs compared to uninfected RBCs, suggesting that gametocytes are more sensitive to alterations in cation concentration than RBCs. Moreover, the addition of 15.6 nM maduramicin enhanced the gametocytocidal activity of the pyrazoleamide PA21A050, which is a promising new antimalarial candidate associated with an increase in intracellular Na⁺ concentration that is proposed to be due to inhibition of PfATP4, a putative Na⁺ pump. These results underscore the importance of cation homeostasis in sexual as well as asexual intraerythrocytic-stage P. falciparum parasites and the potential of targeting this pathway for drug development.     

Gamma interferon is spontaneously released by alveolar macrophages and lung T lymphocytes in patients with pulmonary sarcoidosis.

Gamma interferon (IFN gamma) is a potent immune mediator that plays a central role in enhancing cellular immune processes. This study demonstrates that while lung mononuclear cells from normal individuals spontaneously release little or no interferon (less than 10 U/10(6) cells per 24 h), those from patients with pulmonary sarcoidosis spontaneously release considerable amounts (65 +/- 20 U/10(6) cells per 24 h, P less than 0.02 compared to normals). Furthermore, cells from patients with active disease release far more interferon than those from patients with inactive disease (101 +/- 36 compared to 24 +/- 8 U/10(6) cells per 24 h, P less than 0.02). Characterization of this interferon using acid sensitivity, specific antibody inhibition, and target cell specificity criteria demonstrated that it was almost entirely IFN gamma. This spontaneous release of IFN gamma appeared to be compartmentalized to the lung of these patients in that their blood mononuclear cells spontaneously released little or no IFN gamma (P less than 0.02, compared to sarcoidosis lung mononuclear cells) and no IFN gamma was detected in their serum. Both lung T lymphocytes and alveolar macrophages contributed to the spontaneous release of IFN gamma by lung mononuclear cells from sarcoid patients; purified preparations of T lymphocytes and alveolar macrophages from these patients spontaneously released similar amounts of IFN gamma (56 +/- 21 and 32 +/- 11 U/10(6) cells per 24 h, respectively, P greater than 0.3). At least one role for IFN gamma in the pathogenesis of sarcoidosis appeared to be related to activation of alveolar macrophages, as alveolar macrophages recovered from patients with active disease spontaneously killed [3H]uridine-labeled tumor cell targets (17.7 +/- 4.5% cytotoxicity compared with 2.8 +/- 0.9% in normals, P less than 0.02) and purified IFN gamma enhanced the ability of alveolar macrophages from sarcoidosis patients with inactive disease to kill similar targets (P less than 0.001, compared to alveolar macrophages cultured in medium alone). Treatment of sarcoid patients with corticosteroids, a therapy known to suppress the activity of the disease, caused a marked reduction in the level of spontaneous IFN gamma release by lung mononuclear cells compared with untreated patients (P less than 0.02), which suggests that the effectiveness of corticosteroid therapy in controlling active pulmonary sarcoidosis may, at least in part, be due to suppression of IFN gamma release.     

Genetic relatedness of levofloxacin-nonsusceptible Streptococcus pneumoniae isolates from North America

We characterized 32 levofloxacin-nonsusceptible Streptococcus pneumoniae (LNSP) isolates obtained from a broad geographic region of North America over a 5-year period by using capsular serotypes, antimicrobial susceptibility profiles, BOX-PCR, multilocus sequence typing (MLST), and pulsed-field gel electrophoresis (PFGE). Sixteen international clones identified by the Pneumococcal Molecular Epidemiology Network also were included for comparison. Fifteen serotypes were represented, with serogroups 6, 9, 14, 19, and 23 accounting for 63% of isolates. Among isolates whose quinolone resistance-determining regions were sequenced, all contained gyrA and parC point mutations. Sixty-three percent were penicillin susceptible, and 84% were erythromycin susceptible. BOX-PCR analysis identified 39 different band patterns among 32 LNSP and 16 international clones and grouped 16 isolates, including 2 international clones, into seven unrelated groups of 2 to 4 isolates each. PFGE analysis identified 35 different band patterns among 32 LNSP and 16 international clones and grouped 21 isolates, including 3 international clones, into eight unrelated groups of 2 to 6 isolates each. MLST performed on 10 isolates identified five allelic profiles and separated 9 isolates into four groups of 2 to 3 isolates each. Overall, each typing method indicated that the LNSP were heterogeneous and that resistance to fluoroquinolones was not closely associated with a particular serotype or with coresistance to other antimicrobial classes and suggests that LNSP have likely arisen through independent mutational events as a result of selective pressure. However, seven LNSP were found to be related to three international clones by PFGE.     

Localized eosinophil degranulation mediates disease in tropical pulmonary eosinophilia

To explore the mechanisms underlying the eosinophil-mediated inflammation of tropical pulmonary eosinophilia (TPE), bronchoalveolar lavage (BAL) fluid, serum, and supernatants from pulmonary and blood leukocytes (WBC) from patients with acute TPE (n = 6) were compared with those obtained from healthy uninfected individuals (n = 4) and from patients with asthma (n = 4) or elephantiasis (n = 5). Although there were no significant differences in the levels of interleukin-4 (IL-4), IL-5, IL-13, eotaxin, granulocyte-macrophage colony-stimulating factor, RANTES, or eosinophil cationic protein, there was a marked increase in eosinophil-derived neurotoxin (EDN) both systemically and in the lungs of individuals with TPE compared to each of the control groups (P < 0.02). Moreover, there was a compartmentalization of this response, with EDN levels being higher in the BAL fluid than in the serum (P < 0.02). Supernatants from WBC from either whole blood or BAL cells were examined for chemokines, cytokines, eosinophil degranulation products, and arachidonic acid metabolites. Of the many mediators examined-particularly those associated with eosinophil trafficking-only EDN (in BAL fluid and WBC) and MIP-1alpha (in WBC) levels were higher for TPE patients than for the non-TPE control groups (P < 0.02). These data suggest it is the eosinophilic granular protein EDN, an RNase capable of damaging the lung epithelium, that plays the most important role in the pathogenesis of TPE.     

Intranasal vaccination with a defined attenuated Francisella novicida strain induces gamma interferon-dependent antibody-mediated protection against tularemia

Francisella tularensis is an intracellular gram-negative bacterium that is the causative agent of tularemia and a potential bioweapon. We have characterized the efficacy of a defined F. novicida mutant (DeltaiglC) as a live attenuated vaccine against subsequent intranasal challenge with the wild-type organism. Animals primed with the F. novicida DeltaiglC (KKF24) mutant induced robust splenic gamma interferon (IFN-gamma) and interleukin-12 (IL-12) recall responses with negligible IL-4 production as well as the production of antigen-specific serum immunoglobulin G1 (IgG1) and IgG2a antibodies. BALB/c mice vaccinated intranasally (i.n.) with KKF24 and subsequently challenged with wild-type F. novicida (100 and 1,000 50% lethal doses) were highly protected (83% and 50% survival, respectively) from the lethal challenges. The protection conferred by KKF24 vaccination was shown to be highly dependent on endogenous IFN-gamma production and also was mediated by antibodies that could be adoptively transferred to naive B-cell-deficient mice by inoculation of immune sera. Collectively, the results demonstrate that i.n. vaccination with KKF24 induces a vigorous Th1-type cytokine and antibody response that is protective against subsequent i.n. challenge with the wild-type strain. This is the first report of a defined live attenuated strain providing protection against the inhalation of F. novicida.     

Methamphetamine treatment causes delayed decrease in novelty-induced locomotor activity in mice

Methamphetamine (METH) is a psychostimulant that causes damage to dopamine (DA) axons and to non-monoaminergic neurons in the brain. The aim of the present study was to investigate short- and long-term effects of neurotoxic METH treatment on novelty-induced locomotor activity in mice. Male BALB/c mice, 12–14 weeks old, were injected with saline or METH (i.p., 7.5 mg/kg × 4 times, every 2 h). Behavior and neurotoxic effects were assessed at 10 days, 3 and 5 months following drug treatment. METH administration caused marked decreases in DA levels in the mouse striatum and cortex at 10 days post-drug. However, METH did not induce any changes in novelty-induced locomotor activity. At 3 and 5 months after treatment METH-exposed mice showed significant recovery of DA levels in the striatum and cortex. In contrast, these animals demonstrated significant decreases in locomotor activity at 5 months in comparison to aged-matched control mice. Further assessment of METH toxicity using TUNEL staining showed that the drug induced increased cell death in the striatum and cortex at 3 days after administration. Taken together, these data suggest that delayed deficits in novelty-induced locomotor activity observed in METH-exposed animals are not due to neurodegeneration of DA terminals but to combined effects of METH and age-dependent dysfunction of non-DA intrinsic striatal and/or corticostriatal neurons.