Phosphorus cycling and partitioning in an oligotrophic Everglades wetland ecosystem: a radioisotope tracing study

1. Our goal was to quantify short‐term phosphorus (P) partitioning and identify the ecosystem components important to P cycling in wetland ecosystems. To do this, we added P radiotracer to oligotrophic, P‐limited Everglades marshes. ³²PO₄ was added to the water column in six 1‐m² enclosed mesocosms located in long‐hydroperiod marshes of Shark River Slough, Everglades National Park. Ecosystem components were then repeatedly sampled over 18 days. 2. Water column particulates (>0.45 μm) incorporated radiotracer within the first minute after dosing and stored 95–99% of total water column ³²P activity throughout the study. Soluble (<0.45 μm) ³²P in the water column, in contrast, was always <5% of the ³²P in surface water. Periphyton, both floating and attached to emergent macrophytes, had the highest specific activity of ³²P (Bq g^?131P) among the different ecosystem components. Fish and aquatic macroinvertebrates also had high affinity for P, whereas emergent macrophytes, soil and flocculent detrital organic matter (floc) had the lowest specific activities of radiotracer. 3. Within the calcareous, floating periphyton mats, 81% of the initial ³²P uptake was associated with Ca, but most of this ³²P entered and remained within the organic pool (Ca‐associated = 14% of total) after 1 day. In the floc layer, ³²P rapidly entered the microbial pool and the labile fraction was negligible for most of the study. 4. Budgeting of the radiotracer indicated that ³²P moved from particulates in the water column to periphyton and floc and then to the floc and soil over the course of the 18 day incubations. Floc (35% of total) and soil (27%) dominated ³²P storage after 18 days, with floating periphyton (12%) and surface water (10%) holding smaller proportions of total ecosystem ³²P. 5. To summarise, oligotrophic Everglades marshes exhibited rapid uptake and retention of labile ³²P. Components dominated by microbes appear to control short‐term P cycling in this oligotrophic ecosystem.     

Two soluble glycosyltransferases glycosylate less efficiently in vivo than their membrane bound counterparts

Many Golgi glycosyltransferases are type II membrane proteins which are cleaved to produce soluble forms that are released from cells. Cho and Cummings recently reported that a soluble form of alpha1, 3- galactosyltransferase was comparable to its membrane bound counterpart in its ability to galactosylate newly synthesized glycoproteins (Cho,S.K. and Cummings,R.D. (1997) J. Biol. Chem., 272, 13622-13628). To test the generality of their findings, we compared the activities of the full length and soluble forms of two such glycosyltransferases, ss1,4 N-Acetylgalactosaminyltransferase (GM2/GD2/ GA2 synthase; GalNAcT) and beta galactoside alpha2,6 sialyltransferase (alpha2,6-ST; ST6Gal I), for production of their glycoconjugate products in vivo . Unlike the full length form of GalNAcT which produced ganglioside GM2 in transfected cells, soluble GalNAcT did not produce detectable GM2 in vivo even though it possessed in vitro GalNAcT activity comparable to that of full length GalNAcT. When compared with cells expressing full length alpha2,6-ST, cells expressing a soluble form of alpha2,6-ST contained 3-fold higher alpha2,6-ST mRNA levels and secreted 7-fold greater alpha2,6-ST activity as measured in vitro , but in striking contrast contained 2- to 4-fold less of the alpha2,6-linked sialic acid moiety in cellular glycoproteins in vivo . In summary these results suggest that unlike alpha1,3-galactosyltransferase the soluble forms of these two glycosyltransferases are less efficient at glycosylation of membrane proteins and lipids in vivo than their membrane bound counterparts.      

The early adaptive evolution of calmodulin

Interaction between gene duplication and natural selection in molecular evolution was investigated utilizing a phylogenetic tree constructed by the parsimony procedure from amino acid sequences of 50 calmodulin- family protein members. The 50 sequences, belonging to seven protein lineages related by gene duplication (calmodulin itself, troponin-C, alkali and regulatory light chains of myosin, parvalbumin, intestinal calcium-binding protein, and glial S-100 phenylalanine-rich protein), came from a wide range of eukaryotic taxa and yielded a denser tree (more branch points within each lineage) than in earlier studies. Evidence obtained from the reconstructed pattern of base substitutions and deletions in these ancestral loci suggests that, during the early history of the family, selection acted as a transforming force on expressed genes among the duplicates to encode molecular sites with new or modified functions. In later stages of descent, however, selection was a conserving force that preserved the structures of many coadapted functional sites. Each branch of the family was found to have a unique average tempo of evolutionary change, apparently regulated through functional constraints. Proteins whose functions dictate multiple interaction with several other macromolecules evolved more slowly than those which display fewer protein-protein and protein-ion interactions, e.g., calmodulin and next troponin-C evolved at the slowest average rates, whereas parvalbumin evolved at the fastest. The history of all lineages, however, appears to be characterized by rapid rates of evolutionary change in earlier periods, followed by slower rates in more recent periods. A particularly sharp contrast between such fast and slow rates is found in the evolution of calmodulin, whose rate of change in earlier eukaryotes was manyfold faster than the average rate over the past 1 billion years. In fact, the amino acid replacements in the nascent calmodulin lineage occurred at residue positions that in extant metazoans are largely invariable, lending further support to the Darwinian hypothesis that natural selection is both a creative and a conserving force in molecular evolution.      

Cultured endothelial cells produce heparinlike inhibitor of smooth muscle cell growth

Using cultured cells from bovine and rat aortas, we have examined the possibility that endothelial cells might regulate the growth of vascular smooth muscle cells. Conditioned medium from confluent bovine aortic endothelial cells inhibited the proliferation of growth-arrested smooth muscle cells. Conditioned medium from exponential endothelial cells, and from exponential or confluent smooth muscle cells and fibroblasts, did not inhibit smooth muscle cell growth. Conditioned medium from confluent endothelial cells did not inhibit the growth of endothelial cells or fibroblasts. In addition to the apparent specificity of both the producer and target cell, the inhibitory activity was heat stable and not affected by proteases. It was sensitive flavobacterium heparinase but not to hyaluronidase or chondroitin sulfate ABC lyase. It thus appears to be a heparinlike substance. Two other lines of evidence support this conclusion. First, a crude isolate of glycosaminoglycans (TCA-soluble, ethanol-precipitable material) from endothelial cell-conditioned medium reconstituted in 20 percent serum inhibited smooth muscle cell growth; glycosaminoglycans isolated from unconditioned medium (i.e., 0.4 percent serum) had no effect on smooth muscle cell growth. No inhibition was seen if the glycosaminoglycan preparation was treated with heparinase. Second, exogenous heparin, heparin sulfate, chondroitin sulfate B (dermatan sulfate), chondroitin sulfate ABC, and hyaluronic acid were added to 20 percent serum and tested for their ability to inhibit smooth muscle cell growth. Heparin inhibited growth at concentrations as low as 10 ng/ml. Other glycosaminoglycans had no effect at doses up to 10 μg/ml. Anticoagulant and non- anticoagulant heparin were equally effective at inhibiting smooth muscle cell growth, as they were in vivo following endothelial injury (Clowes and Karnovsk. Nature (Lond.). 265:625-626, 1977; Guyton et al. Circ. Res. 46:625-634, 1980), and in vitro following exposure of smooth muscle cells to platelet extract (Hoover et al. Circ. Res. 47:578-583, 1980). We suggest that vascular endothelial cells may secrete a heparinlike substance in vivo which may regulate the growth of underlying smooth muscle cells.     

Hip joint replacement surgery for idiopathic osteoarthritis aggregates in families

In order to determine whether there is a genetic component to hip or knee joint failure due to idiopathic osteoarthritis (OA), we invited patients (probands) undergoing hip or knee arthroplasty for management of idiopathic OA to provide detailed family histories regarding the prevalence of idiopathic OA requiring joint replacement in their siblings. We also invited their spouses to provide detailed family histories about their siblings to serve as a control group. In the probands, we confirmed the diagnosis of idiopathic OA using American College of Rheumatology criteria. The cohorts included the siblings of 635 probands undergoing total hip replacement, the siblings of 486 probands undergoing total knee replacement, and the siblings of 787 spouses. We compared the prevalence of arthroplasty for idiopathic OA among the siblings of the probands with that among the siblings of the spouses, and we used logistic regression to identify independent risk factors for hip and knee arthroplasty in the siblings. Familial aggregation for hip arthroplasty, but not for knee arthroplasty, was observed after controlling for age and sex, suggesting a genetic contribution to end-stage hip OA but not to end-stage knee OA. We conclude that attempts to identify genes that predispose to idiopathic OA resulting in joint failure are more likely to be successful in patients with hip OA than in those with knee OA.     

Effects of reducing sample size on density estimates of citrus rust mite (Acari: Eriophyidae) on citrus fruit: simulated sampling

The consequence of reducing sample size on the accuracy and precision of estimates of citrus rust mite, Phyllocoptruta oleivora (Ashmead), densities on oranges was investigated. The sample unit was a 1-cm2 surface area on fruit. Sampling plans consisting of 360, 300, 200, 160, 80, 48, 36, or 20 samples per 4 ha were evaluated through computer simulations by using real count data from 32 data sets of 600 sample units per 4 ha. The original and reduced sampling plans were hierarchical with different numbers of sample areas per 4 ha, trees per area, fruit per tree, and samples per fruit. Individual estimates (n=100 simulations per data set) using each plan were sometimes considerably below or above target densities. In an original set of count data with a mean of six mites per cm2, simulations of 36 samples per 4 ha produced individual estimates ranging from one to 16 mites per cm2, whereas 80 samples per 4 ha produced estimates ranging from two to 10 mites per cm2. The plans consisting of 36 or more samples were projected to provide precision levels of 0.25 (SEM/mean) or better at densities of five or more mites per cm2 based on log-data, a projection that needs to be verified under real-grove situations. Each plan consistently provided mite detection in these sampling simulations except those consisting of 20 or 36 samples, which sometimes failed to detect mites when the target density was less than five mites per cm2. The study provided insight into the probable precision, accuracy and detection thresholds for eight candidate sampling plans varying from relatively low to high resource input.     

Biology of three species of Agistemus (Acari: Stigmaeidae): life table parameters using eggs of Panonychus citri or pollen of Malephora crocea as food

The biology and life table parameters of Agistemus industani Gonzalez, A. cyprius Gonzalez, and A. floridanus Gonzalez (Acari: Stigmaeidae) were studied under laboratory conditions using two food sources: Panonychus citri (McGregor) eggs or ice plant, Malephora crocea (Jacquin) Schwantes pollen at 25 degrees C. The larval, protonymph, deutonymph, and adult stages of A. industani fed on citrus red mite eggs. All active stages of A. industani, except the larva, fed on all P. citri stages and the larval stage could not feed on P. citri adults. All immature stages of A. industani fed on M. crocea pollen. Agistemus cyprius larvae fed on P. citri eggs and larvae or ice plant pollen. The nymphal stages fed on P. citri eggs, larvae, and protonymphs but not deutonymphs or adults while A. cyprius deutonymphs and adults fed on all P. citri stages. Adult and nymphal stages of A. cyprius fed on ice plant pollen and successfully completed their development while A. floridanus did not. Agistemus floridanus larvae fed only on P. citri eggs, while the other stages fed on P. citri eggs, larvae, and protonymphs. The developmental times from egg to adult for A. industani and A. cyprius when fed M. crocea pollen were 11.3 and 13.4 days, respectively. Agistemus floridanus was unable to complete its life cycle on a diet of only M. crocea pollen. Agistemus industani, A. cyprius, and A. floridanus completed development from egg to adult in 11.7, 13.8, and 10.8 days, respectively, when fed P. citri eggs. The intrinsic rate of increase (r(m)) values for A. cyprius and A. industani were 0.0311 and 0.1201 per day on the pollen diet. The net reproductive rate (Ro) was 3.58 for A. cyprius and 10.07 for A. industani with generation times (T) of 45.2 and 35.1 days, respectively, on the ice plant pollen diet. The r(m) values for A. cyprius, A. floridanus, and A. industani on the P. citri egg only diet were: 0.0562, 0.1001, and 0.1031 per day, respectively. The Ro values for each species fed P. citri eggs only were: 6.36, 7.90, and 18.70 for A. cyprius, A. floridanus, and A. industani and the generation times (T) for each of the three species were: 35.2, 29.9 and 37.8 days, respectively.     

Mitochondrial DNA and RAPD polymorphisms in the haploid mite Brevipalpus phoenicis (Acari: Tenuipalpidae)

Brevipalpus phoenicis (Geijskes) (Acari: Tenuipalpidae) is recognized as the vector of citrus leprosis virus that is a significant problem in several South American countries. Citrus leprosis has been reported from Florida in the past but no longer occurs on citrus in North America. The disease was recently reported in Central America, suggesting that B. phoenicis constitutes a potential threat to the citrus industries of North America and the Caribbean. Besides B. phoenicis, B. obovatus Donnadieu, and B. californicus (Banks) have been incriminated as vectors of citrus leprosis virus and each species has hundreds of host plants. In this study, Brevipalpus mite specimens were collected from different plants, especially citrus, in the States of Florida (USA) and São Paulo (Brazil), and reared on citrus fruit under standard laboratory conditions. Mites were taken from these colonies for DNA extraction and for morphological species identification. One hundred and two Random Amplified Polymorphic DNA (RAPD) markers were scored along with amplification and sequencing of a mitochondrial cytochrome oxidase subunit I gene fragment (374 bp). Variability among the colonies was detected with consistent congruence between both molecular data sets. The mites from the Florida and Brazilian colonies were morphologically identified as belonging to B. phoenicis, and comprise a monophyletic group. These colonies could be further diagnosed and subdivided geographically by mitochondrial DNA analysis.