Human rhinovirus-induced inflammatory responses are inhibited by phosphatidylserine containing liposomes.

Human rhinovirus (HRV) infections are major contributors to the healthcare burden associated with acute exacerbations of chronic airway disease, such as chronic obstructive pulmonary disease and asthma. Cellular responses to HRV are mediated through pattern recognition receptors that may in part signal from membrane microdomains. We previously found Toll-like receptor signaling is reduced, by targeting membrane microdomains with a specific liposomal phosphatidylserine species, 1-stearoyl-2-arachidonoyl-sn-glycero-3-phospho-L-serine (SAPS). Here we explored the ability of this approach to target a clinically important pathogen. We determined the biochemical and biophysical properties and stability of SAPS liposomes and studied their ability to modulate rhinovirus-induced inflammation, measured by cytokine production, and rhinovirus replication in both immortalized and normal primary bronchial epithelial cells. SAPS liposomes rapidly partitioned throughout the plasma membrane and internal cellular membranes of epithelial cells. Uptake of liposomes did not cause cell death, but was associated with markedly reduced inflammatory responses to rhinovirus, at the expense of only modest non-significant increases in viral replication, and without impairment of interferon receptor signaling. Thus using liposomes of phosphatidylserine to target membrane microdomains is a feasible mechanism for modulating rhinovirus-induced signaling, and potentially a prototypic new therapy for viral-mediated inflammation.

Rhinoviral infection and asthma: the detection and management of rhinoviruses by airway epithelial cells.

Human rhinoviruses (HRV) have been linked to the development of childhood asthma and recurrent acute asthma exacerbations throughout life, and contribute considerably to the healthcare and economic burden of this disease. However, the ability of HRV infections to trigger exacerbations, and the link between allergic status and HRV responsiveness, remains incompletely understood. Whilst the receptors on human airway cells that detect and are utilized by most HRV group A and B, but not C serotypes are known, how endosomal pattern recognition receptors (PRRs) detect HRV replication products that are generated within the cytoplasm remains somewhat of an enigma. In this article, we explore a role for autophagy, a cellular homeostatic process that allows the cell to encapsulate its own cytosolic constituents, as the crucial mechanism controlling this process and regulating the innate immune response of airway epithelial cells to viral infection. We will also briefly describe some of the recent insights into the immune responses of the airway to HRV, focusing on neutrophilic inflammation that is a potentially unwanted feature of the acute response to viral infection, and the roles of IL-1 and Pellinos in the regulation of responses to HRV.

First Report of Pseudomonas syringae pv. actinidiae the Causal Agent of Bacterial Canker of Kiwifruit on Actinidia arguta Vines in New Zealand.

Actinidia arguta is commercially grown in New Zealand and few other countries; the fruit are sometimes sold as kiwiberry or hardy kiwi. In New Zealand, two biovars of Pseudomonas syringae pv. actinidiae have recently been found to cause bacterial canker on both A. chinensis and A. deliciosa, which produce the yellow and green fleshed kiwifruit, respectively (4). In November 2011, in a commercial orchard in the Bay of Plenty, New Zealand, A. arguta 'Tahi' and 'Rua' showed small angular necrotic leaf spots. About 50% of the vines randomly located throughout the orchard showed symptoms. Canker or shoot dieback were not detected on any of the infected plants. Four strains, labeled 13093 to 13096, were isolated onto King's B medium (KB) from leaves selected from four different plants showing symptoms. These four strains were gram-negative, induced a hypersensitive reaction when infiltrated in tobacco plants, lacked cytochrome c oxidase, arginine dehydrolase, and urease activity, and were unable to hydrolyze esculin, starch, and gelatine, and to induce ice nucleation. When plated on KB, these strains showed the same weak fluorescence associated with some strains of P. syringae pv. actinidiae (4). All these characteristics support identification of the strains as P. syringae pv. actinidiae. Using P. syringae pv. actinidiae-specific primers PsaF1/R2 (2), the expected 280-bp fragment was amplified by PCR from genomic DNA extracted from the four strains. The four amplicons were sequenced (GenBank Accession Nos. KF206138 to 41) and found to be 100% identical to each other and to the corresponding DNA fragment of the pathotype strain, ICMP 9617 (AY342165). A similar conclusion was reached using the duplex PCR targeting the ompP1 and the avrD genes (1); two amplicons of 492 and 226 bp were obtained with each of the four strains as expected for P. syringae pv. actinidiae. The DNA sequence of the 492-bp amplicon (KF206134 to 37) was 100% identical to that of strains of P. syringae pv. actinidiae, such as Psa 10627 (JQ934475.1). Strain 13094 isolated from A. arguta and pathotype strain ICMP 9617 were sprayed at a concentration of 3 × 109 cfu/ml on to the undersides of leaves of three 6- to 8-week-old seedlings of A. chinensis'Hort16A' and three similar seedlings of A. deliciosa 'Bruno.' Those are the conditions under which the pathogenicity of strains of P. syringae pv. actinidiae is usually evaluated (4). After 2 weeks of incubation, small necrotic angular spots were observed on all plants inoculated with 13094 or ICMP 9617 but not on the water-treated control plants. The bacteria isolated from those necrotic spots had the same morphological characteristics on KB as P. syringae pv. actinidiae and gave a 280-bp amplicon after PCR with the PsaF1/R2 primers. Leaves of two rooted cuttings of A. arguta'Tahi' were spray inoculated with strain 13094 at a concentration of 2.7 × 109 cfu/ml or with water. Necrotic spots developed on leaves 1 week after inoculation. No spots developed on the water-treated plants. The bacteria isolated from those necrotic spots had the same morphological characteristics on KB as P. syringae pv. actinidiae and gave a 280-bp amplicon after PCR with the PsaF1/R2 primers. Isolation of P. syringae pv. actinidiae from A. arguta has been reported only once before (3). This is this is the first report of P. syringae pv. actinidiae being isolated from A. arguta vines in New Zealand. This limited outbreak did not lead to any loss of production and since then only very few symptoms have been observed in this particular orchard. References: (1) A. Gallelli et al. J. Plant Pathol 93:425, 2011. (2) J. Rees-Gorge et al. Plant Pathol. 59:453, 2010. (3) K. Ushiyama et al. Ann. Phytopath. Soc. Japan 58:476, 1992. (4) J. L. Vanneste et al. Plant Dis. 97:708, 2013.

Muckle-Wells syndrome without mutation in exon 3 of the NLRP3 gene, identified by evidence of excessive monocyte production of functional interleukin 1ß and rapid response to anakinra.

We report a man with lifelong urticaria, night sweats, arthralgia and lethargy. He had high levels of inflammatory markers and serum amyloid A, but no identifiable mutation in exon 3 of the NLRP3 (NOD-like receptor family, pyrin domain-1 containing 3) gene, and no relevant family history. We found marked production of functional interleukin (IL)-1 by the patient's monocytes at baseline and after stimulation with lipopolysaccharide. The patient made an immediate response to treatment with an IL-1ß receptor antagonist. We propose that this patient has Muckle-Wells syndrome without deafness, occurring de novo. Functional screening for IL-1 production could aid diagnosis in future similar cases.

Absorption, distribution, metabolism and excretion of the cholecystokinin-1 antagonist dexloxiglumide in the dog.

Single oral doses of 14C-dexloxiglumide were rapidly and extensively absorbed in dogs and also eliminated rapidly with a short half-life. Following single intravenous doses, dexloxiglumide was characterised as a drug having a high clearance (30.7 and 27.0 ml/min/kg in males and females respectively), a low volume of distribution (Vss, 0.34 and 0.27 L/kg in males and females respectively) and a moderate systemic availability (about 33%). It was extensively bound to plasma proteins (89%). Dexloxiglumide is mainly cleared by the liver. Its renal clearance was minor. In only the kidney, liver and gastrointestinal tract, were concentrations of 14C generally greater than those in plasma. 14C concentrations generally peaked at 0.25h and declined rapidly during 24h being present only in a few tissues (such as the kidney, liver and gastrointestinal tract) at 24h. Single intravenous or oral doses were mainly excreted in the faeces (77-89%), mostly during 24h. Urine contained up to 7.5% dose. Mean recoveries during 7 days ranged between 93-97%. Biliary excretion of 14C was prominent (64% dose during 24h) in the disposition of 14C which was probably also subjected to some limited enterohepatic circulation. Unchanged dexloxiglumide was the major component in plasma. Urine and faeces contained several 14C-components amongst which unchanged dexloxiglumide was the most important (eg. about 55% dose in faeces). LC-MS/MS of urine and bile extracts showed that dexloxiglumide was metabolised mainly by O-demethylation and by conjugation with glucuronic acid.

Absorption, distribution, metabolism and excretion of the cholecystokinin-1 antagonist dexloxiglumide in the rat.

Single oral doses of 14C-dexloxiglumide were rapidly and extensively absorbed in rats, and eliminated more slowly by females than by males. The respective half-lives were about 4.9 and 2.1 h. Following single intravenous doses, dexloxiglumide was characterised as a drug having a low clearance (6.01 and about 1.96 ml/min/kg in males and females respectively), a moderate volume of distribution (Vss, 0.98 and about 1.1 L/kg in males and females respectively) and a high systemic availability. It was extensively bound to plasma proteins (97%). Dexloxiglumide is mainly cleared by the liver. Its renal clearance was minor. In only the liver and gastrointestinal tract, were concentrations of 14C generally greater than those in plasma. Peak 14C concentrations generally occurred at 1-2 h in males and at 2-4 h in females. Tissue 14C concentrations then declined by severalfold during 24 h although still present in most tissues at 24 h but only in a few tissues (such as the liver and gastrointestinal tract) at 168 h. Decline of 14C was less rapid in the tissues of females than in those of males. Single intravenous or oral doses were mainly excreted in the faeces (87-92%), mostly during 24 h and more slowly from females than from males. Urines contained less than 11% dose. Mean recoveries during 7 days when 14C was not detectable in the carcass except in one female rat ranged between 93-101%. Biliary excretion of 14C was prominent (84-91% dose during 24 h) in the disposition of 14C which was also subjected to facile enterohepatic circulation (74% dose). Metabolite profiles in plasma and selected tissues differed. In the former, unchanged dexloxiglumide was the major component whereas in the latter, a polar component was dominant. Urine, bile and faeces contained several 14C-components amongst which unchanged dexloxiglumide was the most important (eg. up to 63% dose in bile). LC-MS/MS showed that dexloxiglumide was metabolised mainly by hydroxylation in the N-(3-methoxypropyl)pentyl sidechain and by O-demethylation followed by subsequent oxidation of the resulting alcohol to a carboxylic acid.

Carbohydrate intolerance and kidney stones in children in the Goldfields.

Renal stones have been reported as a common finding in Australian Aboriginal children. The stones are predominantly urate in composition. We report on five children with nephrolithiasis from the Goldfields region of Western Australia. All were diagnosed when under 5 years of age, the majority being under 3 years. All five children also had lactose intolerance, and we postulate that carbohydrate malabsorption, together with the ensuing chronic diarrhoea and intraluminal breakdown of sugars by enteric bacteria may result in a situation of chronic metabolic acidosis. Chronic metabolic acidosis can lead to protein catabolism, increased urate excretion and the formation of renal stones. Carbohydrate intolerance may be an aetiological factor in the development of renal stones and possibly chronic renal disease, particularly in Aboriginal Australians. Renal disease represents one of the most significant factors affecting the health of Australian Aboriginal people. The incidence of end stage renal failure in this population exceeds that of non-Aboriginals by a factor of 13:1, and this disproportionate figure is increasing. It is likely that chronic renal damage is multifactorial; however, it is probable that at least some aetiological factors have their onset during childhood.

Real-time ultrasonography for determining pregnancy status and viable fetal numbers in ewes.

The reproductive tracts of 26 estrus synchronized, bred ewes were scanned with a portable 5.0 MHz real-time ultrasound unit within 1 to 6 d postbreeding. Intrarectal scanning was performed on alternate days until Days 28 to 30 and twice weekly until Days 50 of gestation. Transabdominal uterine scans were conducted twice weekly from Days 25 to 65 and continued weekly until parturition. A total of 24 ewes (92%) became pregnant. A nonpregnant ewe was recognized 100% of the time by both methods of ultrasonic screening. Correct identification of a gravid ewe as pregnant was 100% from Days 51 to 150 of gestation using transabdominal real-time ultrasonography. There was a significant association (P < 0.005) between the number of lambs born and the number of fetuses observed using transabdominal real-time ultrasonography after Day 25 of gestation. Accurate differentiation of fetal numbers by transabdominal scanning was 100.0% for ewes carrying one lamb and 97.3% for ewes carrying two lambs at Days 51 to 75 of gestation. Fetal attrition was documented in one ewe at Day 49 of gestation. Hydrops allantois was diagnosed in another ewe at 110 d of gestation. A total of 37 lambs were born to 23 ewes in the project flock. No congenital abnormalities were noted in any of the lambs. Transabdominal real-time ultrasonography is a safe, rapid, accurate and practical method for assessing pregnancy status, fetal number and fetal viability in sheep.