Amphibian Pathogens in Southeast Asian Frog Trade

Amphibian trade is known to facilitate the geographic spread of pathogens. Here we assess the health of amphibians traded in Southeast Asia for food or as pets, focusing on Batrachochytrium dendrobatidis (Bd), ranavirus and general clinical condition. Samples were collected from 2,389 individual animals at 51 sites in Lao PDR, Cambodia, Vietnam and Singapore for Bd screening, and 74 animals in Cambodia and Vietnam for ranavirus screening. Bd was found in one frog (n = 347) in Cambodia and 13 in Singapore (n = 419). No Bd was found in Lao PDR (n = 1,126) or Vietnam (n = 497), and no ranavirus was found in Cambodia (n = 70) or Vietnam (n = 4). Mild to severe dermatological lesions were observed in all East Asian bullfrogs Hoplobatrachus rugolosus (n = 497) sampled in farms in Vietnam. Histologic lesions consistent with sepsis were found within the lesions of three frogs and bacterial sepsis in two (n = 4); one had Gram-negative bacilli and one had acid-fast organisms consistent with mycobacterium sp. These results confirm that Bd is currently rare in amphibian trade in Southeast Asia. The presence of Mycobacterium-associated disease in farmed H. rugolosus is a cause for concern, as it may have public health implications and indicates the need for improved biosecurity in amphibian farming and trade.     

Adaptive response of pulmonary arterial smooth muscle to length change.

Hypervasoconstriction is associated with pulmonary hypertension and dysfunction of the pulmonary arterial smooth muscle (PASM) is implicated. However, relatively little is known about the mechanical properties of PASM. Recent advances in our understanding of plastic adaptation in smooth muscle may shed light on the disease mechanism. In this study, we determined whether PASM is capable of adapting to length changes (especially shortening) and regain its contractile force. We examined the time course of length adaptation in PASM in response to step changes in length and to length oscillations mimicking the periodic stretches due to pulsatile arterial pressure. Rings from sheep pulmonary artery were mounted on myograph and stimulated using electrical field stimulation (12-16 s, 20 V, 60 Hz). The length-force relationship was determined at L(ref) to 0.6 L(ref), where L(ref) was a reference length close to the in situ length of PASM. The response to length oscillations was determined at L(ref), after the muscle was subjected to length oscillation of various amplitudes for 200 s at 1.5 Hz. Release (or stretch) of resting PASM from L(ref) to 0.6 (and vice versa) was followed by a significant force recovery (73 and 63%, respectively), characteristic of length adaptation. All recoveries of force followed a monoexponential time course. Length oscillations with amplitudes ranging from 5 to 20% L(ref) caused no significant change in force generation in subsequent contractions. It is concluded that, like many smooth muscles, PASM possesses substantial capability to adapt to changes in length. Under pathological conditions, this could contribute to hypervasoconstriction in pulmonary hypertension.     

Abscisic acid-induced hydrogen peroxide is required for anthocyanin accumulation in leaves of rice seedlings

The role of hydrogen peroxide (H(2)O(2)) in abscisic acid (ABA)-induced anthocyanin accumulation in detached and intact leaves of rice seedlings was investigated. Treatment with ABA resulted in an accumulation of anthocyanins in detached rice leaves. Dimethylthiourea, a chemical trap for H(2)O(2), was observed to be effective in inhibiting ABA-induced accumulation of anthocyanins. Inhibitors of NADPH oxidase (diphenyleneiodonium chloride and imidazole), phosphatidylinositol 3-kinase (wortmannin and LY 294002), and a donor of nitric oxide (N-tert-butyl-alpha-phenylnitrone), which have previously been shown to prevent ABA-induced H(2)O(2) accumulation in detached rice leaves, inhibited ABA-induced anthocyanin increase. Exogenous application of H(2)O(2), however, was found to increase the anthocyanin content of detached rice leaves. In terms of H(2)O(2) accumulation, intact (attached) leaves of rice seedlings of cultivar Taichung Native 1 (TN1) are ABA sensitive and those of cultivar Tainung 67 (TNG67) are ABA insensitive. Upon treatment with ABA, H(2)O(2) and anthocyanins accumulated in leaves of TN1 seedlings but not in leaves of TNG67. Our results, obtained from detached and intact leaves of rice seedlings, suggest that H(2)O(2) is involved in ABA-induced anthocyanin accumulation in this species.     

Osmotic stress-induced changes in cell wall peroxidase activity and hydrogen peroxide level in roots of rice seedlings

The changes in activity of peroxidase (POD) extracted from the cellwalls and the level of H₂O₂ in rice seedling rootstreatedwith mannitol and their correlation with root growth were investigated.Increasing concentrations of mannitol from 92 to 276 mM, which isiso-osmotic with 50 to 150 mM NaCl, progressively reduced rootgrowth and increased POD activities extracted from the cell walls of riceroots.The reduction of growth was also correlated with an increase inH₂O₂ level. Both diamine oxidase (DAO) and NADHperoxidase(NADH-POD) are known to be responsible for the generation ofH₂O₂. Mannitol treatment increased DAO but not NADH-PODactivities in roots of rice seedlings, suggesting that DAO contributes to thegeneration of H₂O₂ in the cell walls of mannitol-treatedroots. An increase in the level of H₂O₂ and the activityof POD extracted from the cell walls of rice roots preceded root growthreduction caused by mannitol. An increase in DAO activity coincided with anincrease in H₂O₂ in roots caused by mannitol. Since DAOcatalyses the oxidation of putrescine, the demonstration that mannitolincreasesthe activity of DAO in roots is consistent with those that mannitol decreasesthe level of putrescine. In conclusion, cell-wall stiffening catalysed by PODispossibly involved in the regulation of root growth reduction caused bymannitol.     

Genomic and proteomic perspectives in cell culture engineering.

In the last few years, the number of biologics produced by mammalian cells have been steadily increasing. The advances in cell culture engineering science have contributed significantly to this increase. A common path of product and process development has emerged in the last decade and the host cell lines frequently used have converged to only a few. Selection of cell clones, their adaptation to a desired growth environment, and improving their productivity has been key to developing a new process. However, the fundamental understanding of changes during the selection and adaptation process is still lacking. Some cells may undergo irreversible alteration at the genome level, some may exhibit changes in their gene expression pattern, while others may incur neither genetic reconstruction nor gene expression changes, but only modulation of various fluxes by changing nutrient/metabolite concentrations and enzyme activities. It is likely that the selection of cell clones and their adaptation to various culture conditions may involve alterations not only in cellular machinery directly related to the selected marker or adapted behavior, but also those which may or may not be essential for selection or adaptation. The genomic and proteomic research tools enable one to globally survey the alterations at mRNA and protein levels and to unveil their regulation. Undoubtedly, a better understanding of these cellular processes at the molecular level will lead to a better strategy for 'designing' producing cells. Herein the genomic and proteomic tools are briefly reviewed and their impact on cell culture engineering is discussed.     

4:2:1 conduction of an AF initiating trigger

A 44 year old male with idiopathic dilated cardiomyopathy was undergoing persistent atrial fibrillation (AF) ablation. Following antral ablation, AF terminated into a regular narrow complex rhythm. Earliest activation was mapped to a focus in the superior vena cava (SVC) which was conducted in a 2:1 ratio to the atria which in turn was conducted with 2:1 ratio to the ventricles, resulting in an unusual 4:2:1 conduction of the SVC tachycardia. 1:1 conduction of the SVC tachycardia to the atrium preceded initiation of AF. During AF, SVC tachycardia continued unperturbed. Sinus rhythm was restored following catheter ablation of the focus.     

Senescence of rice leaves XXXI. Changes of chlorophyll,protein, and polyamine contents and ethylene production during senescence of a chlorophyll-deficient mutant

Leaf senescence of a chlorophylldeficient rice mutant (LT-8) was investigated. At 10 days after planting, the chlorophyll level in the third leaves of rice seedlings of the mutant was about one half that of normal leaves (Norin no. 8), whereas no difference in the protein level could be detected in the two genotypes. The protein level in leaves decreased with increasing age, and no significant difference could be detected during senescence in the two genotypes. Chlorophyll level in the normal leaves also decreased with increasing age. However, the chlorophyll level in the mutant leaves began to decrease only after more than 60% of the initial protein had been degraded. The pattern of ethylene production in the normal leaves was, in general, similar to that in the mutant leaves. Ethylene production first decreased with age, increased to a maximum at day 18, and decreased thereafter. Both spermidine and spermine levels in the leaves of the two genotypes decreased with increasing age. The pattern of the putrescine level in the normal leaves behaved somewhat similar to that in the mutant leaves. However, during the course of senescence, the putrescine level in the mutant leaves was always higher than that in the normal leaves. The possible relationship between endogenous polyamine levels and ethylene production is discussed.     

Exchange of ATP for ADP on high-force cross-bridges of skinned rabbit muscle fibers

The contractile properties of rabbit skinned muscle fibers were studied at 1-2 degrees C in different concentrations of MgATP and MgADP. Double-reciprocal plots of maximum velocity against MgATP concentration at different MgADP concentrations all extrapolated to the same value. This finding suggests that MgATP and MgADP compete for the same site on the cross-bridge, and that the exchange of MgATP for MgADP occurs without a detectable step intervening. The K(m) for ATP was 0.32 mM. The K(i) for MgADP was 0.33 mM. Control experiments suggested that the tortuosity of diffusion paths within the fibers reduced the radial diffusion coefficients for reactants about sixfold. Increasing MgADP from 0.18 to 2 mM at 5 mM ATP or lowering MgATP from 10 to 2 mM at 0.18 mM MgADP, respectively, increased isometric force by 25% and 23%, increased stiffness by 10% and 20%, and decreased maximum velocity by 35% and 31%. Two mechanisms appeared to be responsible. One detained bridges in high-force states, where they recovered from a length step with a slower time course. The other increased the fraction of attached bridges without altering the kinetics of their responses, possibly by an increased activation resulting from cooperative effects of the detained, high-force bridges. The rigor bridge was more effective than the ADP-bound bridge in increasing the number of attached bridges with unaltered kinetics.     

Mutagenicity and polycyclic aromatic hydrocarbon content of fumes from heated cooking oils produced in Taiwan

According to epidemiologic studies, exposure of women to fumes from cooking oils appears to be an important risk factor for lung cancer. Fume samples from three different commercial cooking oils frequently used in Taiwan were collected and analyzed for mutagenicity in the Salmonella/microsome assay. Polycyclic aromatic hydrocarbons were extracted from the samples and identified by HPLC chromatography. Extracts from three cooking oil fumes were found to be mutagenic in the presence of S9 mix. All samples contained dibenz[a,h]anthracene (DB[a,h]A) and benz[a]anthracene (B[a]A). Concentration of DB[a,h]A and B[a]A were 1.9 and 2.2 μg/m³ in fumes from lard oil, 2.1 and 2.3 μg/m³ in soybean oil, 1.8 and 1.3 μg/m³ in peanut oil, respectively. Benzo[a]pyrene (B[a]P) was identified in fume samples of soybean and peanut oil, in concentrations of 19.6 and 18.3 μg/m³, in this order. These results provide experimental evidence and support the findings of epidemiologic observations, in which women exposed to the emitted fumes of cooking oils are at increased risk of contracting lung cancer.     

Ammonium accumulation is associated with senescence of rice leaves

The relationship between ammonium accumulation and senescence of detached rice leaves was investigated. Ammonium accumulation in detached rice leaves coincided closely with dark-induced senescence. Exogenous NH4Cl and methionine sulfoximine, which caused an accumulation of ammonium in detached rice leaves, promoted senescence. Treatments such as light and benzyladenine, which retarded senescence, decreased ammonium level in detached rice leaves. Abscisic acid, which promoted senescence, increased ammonium level in detached rice leaves. The current results suggest that ammonium accumulation may be involved in regulating senescence. Evidence was presented to show that ammonium accumulated in detached rice leaves increases tissue sensitivity to ethylene. The accumulation of ammonium in detached rice leaves during dark-induced senescence is attributed to a decrease in glutamine synthetase activity and an increase in reduction of nitrate.