Shigellosis in Bay of Bengal Islands,India: clinical and seasonal patterns,surveillance of antibiotic susceptibility patterns,and molecular characterization of multidrug-resistant Shigella strains isolated during a 6-year period from 2006 to 2011

This study aims to determine the clinical features and seasonal patterns associated with shigellosis, the antimicrobial resistance frequencies of the isolates obtained during the period 2006–2012 for 22 antibiotics, and the molecular characterization of multidrug-resistant strains isolated from endemic cases of shigellosis in the remote islands of India, with special reference to fluoroquinolone and third-generation cephalosporins resistance. During the period from January 2006 to December 2011, stool samples were obtained and processed to isolate Shigella spp. The isolates were evaluated with respect to their antibiotic resistance pattern and various multidrug resistance determinants, including resistance genes, quinolone resistance determinants, and extended-spectrum β-lactamase (ESBL) production. Morbidity for shigellosis was found to be 9.3 % among children in these islands. Cases of shigellosis occurred mainly during the rainy seasons and were found to be higher in the age group 2–5 years. A wide spectrum of resistance was observed among the Shigella strains, and more than 50 % of the isolates were multidrug-resistant. The development of multidrug-resistant strains was found to be associated with various drug-resistant genes, multiple mutations in the quinolone resistance-determining region (QRDR), and the presence of plasmid-mediated quinolone-resistant determinants and efflux pump mediators. This report represents the first presentation of the results of long-term surveillance and molecular characterization concerning antimicrobial resistances in clinical Shigella strains in these islands. Information gathered as part of the investigations will be instrumental in identifying emerging antimicrobial resistance, for developing treatment guidelines appropriate for that community, and to provide baseline data with which to compare outbreak strains in the future.     

IL-4/IL-13-Dependent Alternative Activation of Macrophages but Not Microglial Cells Is Associated with Uncontrolled Cerebral Cryptococcosis

Both interleukin (IL)-4- and IL-13-dependent Th2-mediated immune mechanisms exacerbate murine Cryptococcus neoformans-induced bronchopulmonary disease. To study the roles of IL-4 and IL-13 in cerebral cryptococcosis, IL-4 receptor α-deficient (IL-4Rα^−/−), IL-4-deficient (IL-4^−/−), IL-13-deficient (IL-13^−/−), IL-13 transgenic (IL-13^T/+), and wild-type mice were infected intranasally. IL-13^T/+ mice displayed a higher fungal brain burden than wild-type mice, whereas the brain burdens of IL-4Rα^−/−, IL-4^−/−, and IL-13^−/− mice were significantly lower as compared with wild-type mice. On infection, 68% of wild-type mice and 88% of IL-13-overexpressing IL-13^T/+ mice developed significant cerebral lesions. In contrast, only a few IL-4Rα^−/−, IL-4^−/−, and IL-13^−/− mice had small lesions in their brains. Furthermore, IL-13^T/+ mice harbored large pseudocystic lesions in the central nervous system parenchyma, bordered by voluminous foamy alternatively activated macrophages (aaMphs) that contained intracellular cryptococci, without significant microglial activation. In wild-type mice, aaMphs tightly bordered pseudocystic lesions as well, and these mice, in addition, showed microglial cell activation. Interestingly, in resistant IL-4^−/−, IL-13^−/−, and IL-4Rα^−/− mice, no aaMphs were discernible. Microglial cells of all mouse genotypes neither internalized cryptococci nor expressed markers of alternative activation, although they displayed similar IL-4Rα expression levels as macrophages. These data provide the first evidence of the development of aaMphs in a central nervous system infectious disease model, pointing to distinct roles of macrophages versus microglial cells in the central nervous system immune response against C. neoformans.The opportunistic pathogenic yeast Cryptococcus neoformans causes life-threatening fungal infections of most internal organs including the central nervous system (CNS), primarily in patients affected by immunodeficiency syndromes such as AIDS.¹ The pathogenesis of cryptococcosis is not fully understood, however, especially in cases of different levels of immunocompetence. It is generally accepted that the fungus first invades the respiratory system, where it leads to relatively mild or asymptomatic bronchopneumonia in the immunocompetent.^2,3,4,5 Fungemia with generalization of the infection may result from reduced immunological control mechanisms.^6,7,8,9 Invasion of the CNS with subsequent development of meningoencephalitis is the major cause of death during cryptococcosis.^10,11The precise reaction pattern of recruited inflammatory cells, especially monocytes/macrophages, due to fungal invasion of the CNS parenchyma has been addressed mainly via analysis of helper T cell (Th)1 responses.¹² In this context, in addition to protective Th1-driven immune responses, the role of Th2 cytokines has gained interest recently.¹³ The major Th2 cytokines interleukin (IL)-4 and IL-13 act via the IL-4Rα chain together with the γ_c chain or the IL-13Rα1/2 chains, and regulate macrophage functional status.¹⁴ IL-4 has been shown to be detrimental in murine models of systemic and pulmonary cryptococcosis,^{6,15,16,17,18} and we have recently illustrated the role of IL-13 in inducing the formation of alternatively activated macrophages (aaMphs) in murine pulmonary cryptococcosis.¹⁹The activation phenotype of macrophages may critically influence the regulatory mechanisms by which inflammation and infection in the CNS are controlled. According to the current paradigm, classically activated macrophages are primed by interferon-γ and produce tumor necrosis factor, IL-1, oxygen and nitrogen radicals,²⁰ thereby producing proinflammatory cytokines that regulate the Th1 immune response. In contrast, aaMph²¹ develop in response to Th2 cytokine stimulation such as IL-4 and IL-13 and are characterized by expression of genes associated with endocytosis and tissue repair such as arginase-1, mannose receptor (CD206), found-in-inflammatory-zone (FIZZ), and chitinase 3-like 3 (YM1) and largely fail to produce nitric oxide (NO) due to their induction of arginase.²² As such, they are thought to be involved in tissue repair and remodeling,^22,23 in protection against diet-induced obesity,^24,25 and schistosomiasis,²⁶ but they may also elicit adverse tissue processes such as pulmonary or liver fibrosis.^{27,28,29,30,31} In particular, their development renders the host vulnerable to infection with pathogens where macrophage activation and killing functions are required.³²In murine models of pulmonary C. neoformans infection, aaMph have been shown to be associated with uncontrolled lung infection.^18,19 The role of aaMph versus classically activated macrophage in the CNS due to pulmonary infection with the neurotropic pathogen C. neoformans has not been defined yet. In this study, we aimed to characterize the morphology and functional status of CNS macrophages in cerebral cryptococcosis following intranasal infection of susceptible wild-type and IL-13-transgenic BALB/c mice. Moreover, using mice unable to produce IL-4 or IL-13 or respond to both (IL-4Rα^−/− mice), we show that abrogation of CNS aaMph development is associated with controlled infection.     

Real-time PCR for diagnosis of human bocavirus infections and phylogenetic analysis

The human bocavirus (hBoV) was first described in 2005 in respiratory tract samples. The clinical relevance of hBoV is still unclear. The aim of our study was to establish a real-time PCR assay for the detection and quantification of hBoV DNA, to apply the real-time assay for the analysis of stool and serum samples for the presence of hBoV DNA, and to perform a phylogenetic analysis of the hBoV positive samples. A total of 834 nasopharyngeal aspirates (NPA), 10 serum samples, and 31 stool samples of children with acute respiratory diseases were retrospectively tested. For phylogenetic analysis, 968 bp of the VP2 gene were sequenced from 69 hBoV-positive NPA samples. The qualitative results of the real-time hBoV PCR were in good agreement with a conventional hBoV PCR. We found that 12% of the NPA were positive for hBoV DNA. The median viral load in the NPA was 4.9 x 10(3) copies/ml (range, 2.7 x 10 degrees to 1.5 x 10(11) copies/ml). There was no difference of the hBoV load in NPA between children with or without known coinfection, but the load was significantly higher in children with bronchitis than in children with the diagnosis of febrile seizures. hBoV DNA was found in 1 of 10 serum samples and in 14 of 31 stool samples. hBoV sequence identity was >99% in the VP2 region. In conclusion, hBoV DNA can be found in NPA samples at very high titers. In addition to being found in the respiratory tract, hBoV was found in stool samples. The clinical relevance of these findings remains to be determined.     

When administered to rats in a cold environment, 3,4-methylenedioxymethamphetamine reduces brown adipose tissue thermogenesis and increases tail blood flow: effects of pretreatment with 5-HT1A and dopamine D2 antagonists

When given in a warm environment MDMA (3,4-methylenedioxymethamphetamine, ecstasy) causes hyperthermia by increasing interscapular brown adipose tissue (iBAT) heat production and decreasing heat loss via cutaneous vasoconstriction. When given in a cold environment, however, MDMA causes hypothermia by an unknown mechanism. This paper addresses these mechanisms and in addition examines whether antagonists at 5-HT(1A) and D(2) receptors reduce the hypothermic action of MDMA. Male Sprague-Dawley rats instrumented with a Doppler probe for measuring tail blood flow, and probes for measuring core and iBAT temperatures, were placed in a temperature-controlled chamber. The chamber temperature was reduced to 10 degrees C and vehicle (0.5 ml Ringer), the 5-HT(1A) antagonist WAY 100635 (0.5 mg/kg), the D(2) antagonist spiperone (20 mug/kg), or the combination of Way 100635 and spiperone were injected s.c. Thirty minutes later the antagonists were injected again along with MDMA (10 mg/kg) or vehicle. MDMA reduced core body temperature by preventing cold-elicited iBAT thermogenesis and by transiently reversing cold-elicited cutaneous vasoconstriction. Pretreatment with WAY 100635 prevented MDMA induced increases in tail blood flow, and briefly attenuated MDMA's effects on iBAT and core temperature. While spiperone alone failed to affect any of the parameters, the combination of spiperone and WAY 100635 decreased MDMA-mediated hypothermia by attenuating both the effects on tail blood flow and iBAT thermogenesis. MDMA's prevention of cold-induced iBAT thermogenesis appears to have a central origin as it rapidly reverses cold-induced increases in iBAT sympathetic nerve discharge in anesthetized rats. Our results demonstrate that MDMA in a cold environment reduces core body temperature by inhibiting iBAT thermogenesis and tail artery vasoconstriction and suggest that mechanisms by which this occurs include the activation of 5-HT1A and dopamine D2 receptors.     

Alterations in cadherin and catenin expression during the biological progression of melanocytic tumours.

AIMS: Compelling evidence from cell culture studies implicates cadherins in the neoplastic progression of melanocytic tumours but few reports describe the expression of cadherins and the related transmembrane proteins, catenins, in a full range of benign and malignant excised melanocytic tumours. METHODS: Using immunohistochemistry and western blotting after tissue fractionation, the pattern of expression of cadherins/catenins was studied in a range of surgically excised melanocytic tumours, from dysplastic naevi to stage III cutaneous metastatic malignant melanoma. RESULTS: Appropriate membranous expression of E-cadherins and P-cadherins is seen in dysplastic naevocytes with an epithelioid phenotype and is largely maintained with malignant transformation to radial growth phase melanoma and primary vertical growth phase malignant melanoma. Loss of membranous E-cadherin is seen in a small number of vertical growth phase melanomas only when metastasis has occurred. However, there is a concomitant dramatic loss of membranous P-cadherin expression in all melanomas at the same stage. A minority of metastatic melanomas show de novo membranous N-cadherin expression in comparison with dysplastic naevi and primary melanoma. Membranous expression of the desmosomal cadherin, desmoglein, was not seen in any tumour studied. Frequently, beta catenin is aberrantly produced in the cytoplasm of cells in dysplastic naevi and metastatic malignant melanoma, with an implied compromise to adhesive function. Furthermore, membranous gamma catenin expression was not seen in any of the 70 melanocytic tumours studied, implying obligatory transmembrane binding of cadherins to beta catenin for maintenance of adhesive function. CONCLUSIONS: The most important alterations in membranous cadherin and catenin expression are seen late in the biological progression of melanocytic tumours at the stage of "in transit" or regional lymph node metastasis, with implications for tumour growth, invasion, and dissemination.     

Dopamine D2 receptor stimulation inhibits cold-initiated thermogenesis in brown adipose tissue in conscious rats

Dopamine D(2)-like receptor agonists cause hypothermia. We investigated whether inhibiting heat production by interscapular brown adipose tissue (iBAT), a major thermogenic organ in rats, contributes to hypothermia caused by dopamine D(2)-like receptor agonists. Temperature of iBAT and tail artery blood flow were measured in conscious rats. Activity in postganglionic sympathetic nerves supplying iBAT was assessed in anesthetized rats. Conscious rats were housed in a warm cage maintained at 26-28 degrees C and then transferred to a cold cage at 5-10 degrees C to induce iBAT thermogenesis. Cold exposure increased iBAT temperature (+0.7+/-0.1 degrees C, 30 min after transferring to the cold cage, P<0.01, n=54). The mixed dopamine D(2)/D(3) receptor agonist, 7-hydroxy-2-(di-n-propylamino)tetralin (7-OH-DPAT, 0.5 mg/kg s.c.) reversed the cold-induced increase in iBAT temperature (-2.8+/-0.2 degrees C at 30 min after 7-OH-DPAT treatment during cold exposure vs. +0.3+/-0.1 degrees C at 30 min after vehicle treatment during cold exposure, n=8). These temperature changes were blocked by pre-treatment with the D(2) receptor antagonists spiperone (20 microg/kg i.p.) and L-741,626 (2.5 mg/kg i.p.), but not by the selective D(3) receptor antagonist SB-277011A (10 mg/kg i.p.). Another mixed dopamine D(2)/D(3) receptor agonist, quinpirole (0.5 mg/kg s.c.) also reversed cold-induced iBAT thermogenesis, and this effect was also prevented by pre-treatment with spiperone, but not with a peripherally acting dopamine receptor antagonist, domperidone (2 mg/kg s.c.). Neither 7-OH-DPAT nor quinpirole reversed cutaneous vasoconstriction elicited by cold exposure. In anesthetized rats, quinpirole (0.5 mg/kg i.v.) abolished iBAT sympathetic nerve discharge elicited by cooling the trunk, and this change was reversed by spiperone (20 microg/kg i.v.). These results demonstrate that activation of CNS dopamine D(2) receptors inhibits sympathetically-mediated iBAT thermogenesis in response to cold exposure. Furthermore, they suggest that in rats hypothermia induced by dopamine D(2) receptor agonists in cold environments is mainly due to decreased heat production rather than to increased heat loss.     

The Subarachnoid Hemorrhage International Trialists (SAHIT) Repository: Advancing Clinical Research in Subarachnoid Hemorrhage

Researchers and other stakeholders continue to express concern about the failure of randomized clinical trials (RCT) in subarachnoid hemorrhage (SAH) to show efficacy of new treatments. Pooled data may be particularly useful to generate hypotheses about causes of poor outcomes and reasons for failure of RCT in SAH, and strategies to improve them. Investigators conducting SAH research collaborated to share data with the intent to develop a large repository of pooled individual patient data for exploratory analysis and testing of new hypotheses relevant to improved trial design and analysis in SAH. This repository currently contains information on 11,443 SAH patients from 14 clinical databases, of which 9 are datasets of recent RCTs and 5 are datasets of prospective observational studies and hospital registries. Most patients were managed in the last 15 years. Data validation and quality checks have been conducted and are satisfactory. Data is available on demographic, clinical, neuroimaging, and laboratory results and various outcome measures. We have compiled the largest known dataset of patients with SAH. The SAHIT repository may be an important resource for advancing clinical research in SAH and will benefit from contributions of additional datasets.     

Raphe region mediates changes in cutaneous vascular tone elicited by stimulation of amygdala and hypothalamus in rabbits

Raphe pallidus/parapyramidal neurons control cutaneous vasoconstriction induced by noxious stimuli. To determine whether they mediate forebrain-induced cutaneous vasoconstriction, we assessed changes in ear pinna blood flow elicited by electrical stimulation of amygdala and hypothalamus before and after injection of muscimol into the raphe/parapyramidal region. We compared ear flow with simultaneously recorded mesenteric flow. Experiments were performed in rabbits anesthetized with urethane (1.25-1.5 g/kg), paralysed and mechanically ventilated. Amygdala stimulation reduced skin conductance from 0.32+/-0.06 to 0.10+/-0.02 cm/s per mmHg (P<0.05, n=9), without effect on mesenteric conductance. Hypothalamic stimulation caused vasoconstriction in both cutaneous and mesenteric beds (conductances fell from 0.27+/-0.05 to 0.05+/-0.02 cm/s per mmHg and from 0.27+/-0.06 to 0.14+/-0.04 cm/s per mmHg (P<0.05, n=9), respectively). Muscimol microinjection (5 nmol in 100 nl) to raphe/parapyramidal region eliminated amygdala- and hypothalamus-induced skin vasoconstriction (pre-stimulus conductance 0.42+/-0.13 and 0.41+/-0.11 cm/s per mmHg, post-stimulus 0.41+/-0.12 and 0.39+/-0.10 cm/s per mmHg, respectively), but not hypothalamically-induced mesenteric vasoconstriction (pre-stimulus 0.29+/-0.06, post-stimulus 0.16+/-0.03 cm/s per mmHg, P<0.05, n=8). The latter was strongly attenuated by bilateral injection of muscimol to the rostral ventrolateral medulla. Data suggest that descending hypothalamo-spinal and amygdala-spinal pathways constricting the cutaneous vascular bed relay in the raphe/parapyramidal area. A relay in the rostral ventrolateral medulla contributes substantially to mesenteric vasoconstriction elicited from the hypothalamus.     

A phase II evaluation of cisplatin and 5-fluorouracil in patients with advanced squamous cell carcinoma of the cervix: a Gynecologic Oncology Group Study

Cisplatin is one of the most active single agents in advanced squamous cell carcinoma of the cervix and is synergistic with 5-fluorouracil in the laboratory. The Gynecologic Oncology Group has conducted a phase II trial in which cisplatin at 50 mg/ml2 given intravenously on Day 1 was combined with 5-fluorouracil 1000 mg/m2 daily given as a 24-hr infusion on Days 1-5. Treatment was repeated every 21 days. Fifty-five patients were treated with this regimen, resulting in seven complete remissions (12.7%) and five partial remissions (9.1%). The median survival was 6.4 months. Toxic effects of grade 2 or greater were leukopenia in 10 patients (18.2%), thrombocytopenia in 2 patients (3.6%), gastrointestinal effects in 25 patients (45.5%), and renal effects in 1 patient (1.8%). On the basis of these results, the cisplatin and 5-fluorouracil regimen does not appear to have any advantage over cisplatin alone in advanced cervical cancer.