Influenza virus inoculum volume is critical to elucidate age‐dependent mortality in mice

The elderly exhibit increased mortality to influenza viral infection for unclear reasons. Mice are frequently used to model how aging impacts disease. Several studies have shown that aged mice exhibit an increased mortality to influenza virus, but two recent studies demonstrated the opposite. These two studies administered the virus intranasally in 20 µL, whereas the other studies used a viral inoculum in at least 30 µL. To determine whether the volume of the inoculum could explain the conflicting reports, we infected young and aged mice via intranasal instillation of 40 µL or 20 µL containing 1 x 10⁴ plaque‐forming units (PFU) of H1N1 influenza virus. We found that intranasal administration of 40 µL but not 20 µL of inoculum resulted in age‐dependent mortality in mice. Compared to aged mice infected with 40 µL inoculum, those infected with 20 µL inoculum showed reduced levels of live virus and IFN‐β in the lung 3 days postinfection. Furthermore, aged mice administered 40 µL of Evans blue intranasally displayed increased dye retention in their bronchoalveolar lavage fluid compared to those administered 20 µL of Evans blue. Our data demonstrate that the inoculating volume of virus is critical for adequate delivery of influenza virus to the lung and thus for efficient infection of aged mice. These findings shed light on discrepant results in the literature regarding aged mice and influenza infection, and establish that mice can be used to examine how aging impacts the response to this biomedically important infection.     

PMR5, an acetylation protein at the intersection of pectin biosynthesis and defense against fungal pathogens

Powdery mildew (Golovinomyces cichoracearum), one of the most prolific obligate biotrophic fungal pathogens worldwide, infects its host by penetrating the plant cell wall without activating the plant's innate immune system. The Arabidopsis mutant powdery mildew resistant 5 (pmr5) carries a mutation in a putative pectin acetyltransferase gene that confers enhanced resistance to powdery mildew. Here, we show that heterologously expressed PMR5 protein transfers acetyl groups from [¹⁴C]‐acetyl‐CoA to oligogalacturonides. Through site‐directed mutagenesis, we show that three amino acids within a highly conserved esterase domain in putative PMR5 orthologs are necessary for PMR5 function. A suppressor screen of mutagenized pmr5 seed selecting for increased powdery mildew susceptibility identified two previously characterized genes affecting the acetylation of plant cell wall polysaccharides, RWA2 and TBR. The rwa2 and tbr mutants also suppress powdery mildew disease resistance in pmr6, a mutant defective in a putative pectate lyase gene. Cell wall analysis of pmr5 and pmr6, and their rwa2 and tbr suppressor mutants, demonstrates minor shifts in cellulose and pectin composition. In direct contrast to their increased powdery mildew resistance, both pmr5 and pmr6 plants are highly susceptibile to multiple strains of the generalist necrotroph Botrytis cinerea, and have decreased camalexin production upon infection with B. cinerea. These results illustrate that cell wall composition is intimately connected to fungal disease resistance and outline a potential route for engineering powdery mildew resistance into susceptible crop species.     

Co-cultivation of the anaerobic fungus Anaeromyces robustus with Methanobacterium bryantii enhances transcription of carbohydrate active enzymes

Journal of Industrial Microbiology & Biotechnology - Anaerobic gut fungi are biomass degraders that form syntrophic associations with other microbes in their native rumen environment. Here,...     

Human Semaphorin 3 Variants Link Melanocortin Circuit Development and Energy Balance

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Yeast homologue of neuronal frequenin is a regulator of phosphatidylinositol-4-OH kinase

In metazoans, certain calmodulin-related calcium-binding proteins (recoverins, neurocalcins and frequenins) are found at highest levels in excitable cells, but their physiological roles are largely uncharacterized. Here we show that Saccharomyces cerevisiae contains a frequenin homologue, Frq1, and that its target is Pik1, a phosphatidylinositol-4-OH kinase. Frq1 binds to a conserved sequence motif in Pik1 outside Pik1's catalytic domain and stimulates its activity in vitro. N-myristoylated Frq1 may also assist in Pik1 localization.     

Research letter: Predictive value of a self-reported history of varicella infection in determining immunity in adults

THE RECENT INTRODUCTION OF A VACCINE FOR VARICELLA has raised questions about whether, for adults, a patient''s history of varicella infection is useful in determining if vaccination is necessary. We report findings on 184 family medicine patients aged 18 to 65 years who were asked if they had a history of varicella infection and were subsequently tested for varicella antibodies. A history of infection was positive for 114 (62%) of the participants and negative or uncertain for 70 (38%). All 114 subjects who reported a varicella infection history were immune. All 4 subjects who were not immune reported an uncertain or negative infection history. Except for people who are at increased risk of varicella infection or complications from infection, serologic testing may not be required for adults in the general population who have a history of varicella infection. Varicella infection (chickenpox) is highly contagious¹ and is spread by respiratory droplets or direct contact. Varicella can occur in nonimmune adults, in whom severity of the infection increases with age,² often causing serious morbidity and loss of work time. In Canada, 70% of 53 reported deaths caused by varicella between 1987 and 1996 occurred in adolescents and adults.³ When acquired during pregnancy, varicella can cause significant maternal, perinatal and infant morbidity.³ After varicella infection, more than 95% of people develop antibodies,⁴ which are detected by serologic testing and indicate lifelong immunity.The recent introduction of a vaccine for varicella means that nonimmune adults may benefit from vaccination.⁵ This has led to studies to determine whether a self-reported history of varicella infection is an accurate indication of the presence of antibodies. Studies to date have yielded disparate conclusions and recommendations,^{6,7,8,9,10,11} and no study has yet investigated this question for adults in a primary care population. Our objective was to determine, in an adult primary care population, the accuracy of a self-reported history of varicella infection in determining immunity.St. Michael''s Hospital Family Practice Unit serves a population with a wide range of socioeconomic groups and countries of origin. The mean patient age is 44 years, and 60% of patients are female. On randomly selected clinic dates between October and December 2000 we enrolled patients aged 18 to 65 years who were having blood taken for reasons other than serologic testing for varicella antibodies. Using a structured interview, patients were asked about their history of varicella infection (Box 1), and their blood samples were analyzed for antibody titres using the VZVscan latex agglutination test (Becton Dickinson, Cockeysville, Md.). We excluded patients with psychiatric or medical conditions impairing memory, a history of varicella vaccination or active varicella infection. All patients gave informed consent, and the study was approved by the St. Michael''s Hospital Research Ethics Board.Open in a separate windowBox 1A sample size of 200 patients provides power to detect a 95% confidence interval (CI) of ± 3% if the baseline positive predictive value of a history of varicella is 95%.Of 204 participants enrolled, 184 had a serologic test for varicella antibodies. For unknown reasons, the serologic test was not performed in the remaining 20 cases.Of the 184 participants whose blood sample was tested for varicella antibodies, 101 (55%) were women, 117 (64%) were born in North America, and 107 (58%) had postsecondary education. The mean age was 43 (standard deviation 12.8) years. Participants for whom serologic testing was not done had similar characteristics.All 114 (62%) of the subjects who reported a history of varicella infection were immune, for a positive predictive value of 100% (95% CI 97%–100%) (Open in a separate windowIn this study, a self-reported history of varicella infection was a highly accurate indicator of immunity to the pathogen. The 98% (180/184) seroprevalence of varicella–zoster virus antibodies in our study population was slightly higher than other population estimates,² and consequently a large majority of those who were uncertain of or reported no past infection were also immune.Although we recruited participants from only 1 family medicine clinic in an urban teaching centre, the patient population of the clinic is diverse in terms of patient country of origin, socioeconomic status and demographic characteristics.Whether particular vaccination strategies are appropriate or cost-effective depends on the population examined and their circumstances. The costs of serologic testing and vaccination and the potential financial, social and medical consequences of infection should be considered.For people who are at increased risk of varicella infection or for whom it is crucial to establish immunity, such as health care workers,^9,10,12,13 pregnant women^7,11,14 and household contacts of immunocompromised people, it may be prudent to have them undergo routine serologic testing regardless of their self-reported infection history.In this primary care setting, a positive history of varicella infection was an accurate indicator of the presence of antibodies. Except for people at high risk of varicella infection, serologic testing may not be required for adults in the general population who have had the infection. Vaccination should be offered to nonimmune patients.     

Mu opioid receptor: a gateway to drug addiction

Mu opioid receptors mediate positive reinforcement following direct (morphine) or indirect (alcohol, cannabinoids, nicotine) activation, and our understanding of mu receptor function is central to the development of addiction therapies. Recent data obtained in native neurons confirm that mu receptor signaling and regulation are strongly agonist-dependent. Current functional mapping reveals morphine-activated neurons in the extended amygdala and early genomic approaches have identified novel mu receptor-associated proteins. A classification of about 30 genes either promoting or counteracting the addictive properties of morphine is proposed from the analysis of knockout mice data. The targeting of effectors or regulatory proteins, beyond the mu receptor itself, might provide valuable strategies to treat addictive disorders.     

Impaired NO signaling in small pulmonary arteries of chronically hypoxic newborn piglets

We performed studies to determine whether chronic hypoxia impairs nitric oxide (NO) signaling in resistance level pulmonary arteries (PAs) of newborn piglets. Piglets were maintained in room air (control) or hypoxia (11% O(2)) for either 3 (shorter exposure) or 10 (longer exposure) days. Responses of PAs to a nonselective NO synthase (NOS) antagonist, N(omega)-nitro-L-arginine methylester (L-NAME), a NOS-2-selective antagonist, aminoguanidine, and 7-nitroindazole, a NOS-1-selective antagonist, were measured. Levels of NOS isoforms and of two proteins involved in NOS signaling, heat shock protein (HSP) 90 and caveolin-1, were assessed in PA homogenates. PAs from all groups constricted to L-NAME but not to aminoguanidine or 7-nitroindazole. The magnitude of constriction to L-NAME was similar for PAs from control and hypoxic piglets of the shorter exposure period but was diminished for PAs from hypoxic compared with control piglets of the longer exposure period. NOS-3, HSP90, and caveolin-1 levels were similar in hypoxic and control PAs. These findings indicate that NOS-3, but not-NOS 2 or NOS-1, is involved with basal NO production in PAs from both control and hypoxic piglets. After 10 days of hypoxia, NO function is impaired in PAs despite preserved levels of NOS-3, HSP90, and caveolin-1. The development of NOS-3 dysfunction in resistance level PAs may contribute to the progression of chronic hypoxia-induced pulmonary hypertension in newborn piglets.     

Arachidonic acid metabolites and an early stage of pulmonary hypertension in chronically hypoxic newborn pigs

Our purpose was to determine whether production of arachidonic acid metabolites, particularly cyclooxygenase (COX) metabolites, is altered in 100-400-microm-diameter pulmonary arteries of piglets at an early stage of pulmonary hypertension. Piglets were raised in either room air (control) or hypoxia for 3 days. A cannulated artery technique was used to measure responses of 100-400-microm-diameter pulmonary arteries to arachidonic acid, a prostacyclin analog, or the thromboxane mimetic. Radioimmunoassay was used to determine pulmonary artery production of thromboxane B(2) (TxB(2)) and 6-keto-prostaglandin F(1alpha) (6-keto-PGF(1alpha)), the stable metabolites of thromboxane and prostacyclin, respectively. Assessment of abundances of COX pathway enzymes in pulmonary arteries was determined by immunoblot technique. Arachidonic acid induced less dilation in pulmonary arteries from hypoxic than in pulmonary arteries from control piglets. Pulmonary artery responses to prostacyclin and were similar for both groups. 6-Keto-PGF(1alpha) production was reduced, whereas TxB(2) production was increased in pulmonary arteries from hypoxic piglets. Abundances of both COX-1 and prostacyclin synthase were reduced, whereas abundances of both COX-2 and thromboxane synthase were unaltered in pulmonary arteries from hypoxic piglets. At least partly due to altered abundances of COX pathway enzymes, a shift in production of arachidonic acid metabolites, away from dilators toward constrictors, may contribute to the early phase of chronic hypoxia-induced pulmonary hypertension in newborn piglets.