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Biomedical subjects

G J Anderson

Publications and source records attributed to G J Anderson.

At least 37 records · Page 2Linked to original sources

Characterization of clinical isolates of Klebsiella pneumoniae from 19 laboratories using the National Committee for Clinical Laboratory Standards extended-spectrum beta-lactamase detection methods.

Extended-spectrum beta-lactamases (ESBLs) are enzymes found in gram-negative bacilli that mediate resistance to extended-spectrum cephalosporins and aztreonam. In 1999, the National Committee for Clinical Laboratory Standards (NCCLS) published methods for screening and confirming the presence of ESBLs in Klebsiella pneumoniae, Klebsiella oxytoca, and Escherichia coli. To evaluate the confirmation protocol, we tested 139 isolates of K. pneumoniae that were sent to Project ICARE (Intensive Care Antimicrobial Resistance Epidemiology) from 19 hospitals in 11 U.S. states. Each isolate met the NCCLS screening criteria for potential ESBL producers (ceftazidime [CAZ] or cefotaxime [CTX] MICs were > or =2 microg/ml for all isolates). Initially, 117 (84%) isolates demonstrated a clavulanic acid (CA) effect by disk diffusion (i.e., an increase in CAZ or CTX zone diameters of > or =5 mm in the presence of CA), and 114 (82%) demonstrated a CA effect by broth microdilution (reduction of CAZ or CTX MICs by > or =3 dilutions). For five isolates, a CA effect could not be determined initially by broth microdilution because of off-scale CAZ results. However, a CA effect was observed in two of these isolates by testing cefepime and cefepime plus CA. The cefoxitin MICs for 23 isolates that failed to show a CA effect by broth microdilution were > or =32 microg/ml, suggesting either the presence of an AmpC-type beta-lactamase or porin changes that could mask a CA effect. By isoelectric focusing (IEF), 7 of the 23 isolates contained a beta-lactamase with a pI of > or =8.3 suggestive of an AmpC-type beta-lactamase; 6 of the 7 isolates were shown by PCR to contain both ampC-type and bla(OXA) genes. The IEF profiles of the remaining 16 isolates showed a variety of beta-lactamase bands, all of which had pIs of < or =7.5. All 16 isolates were negative by PCR with multiple primer sets for ampC-type, bla(OXA), and bla(CTX-M) genes. In summary, 83.5% of the K. pneumoniae isolates that were identified initially as presumptive ESBL producers were positive for a CA effect, while 5.0% contained beta-lactamases that likely masked the CA effect. The remaining 11.5% of the isolates studied contained beta-lactamases that did not demonstrate a CA effect. An algorithm based on phenotypic analyses is suggested for evaluation of such isolates.

Algorithms↗

Cloning and gastrointestinal expression of rat hephaestin: relationship to other iron transport proteins.

The membrane-bound ceruloplasmin homolog hephaestin plays a critical role in intestinal iron absorption. The aims of this study were to clone the rat hephaestin gene and to examine its expression in the gastrointestinal tract in relation to other genes encoding iron transport proteins. The rat hephaestin gene was isolated from intestinal mRNA and was found to encode a protein 96% identical to mouse hephaestin. Analysis by ribonuclease protection assay and Western blotting showed that hephaestin was expressed at high levels throughout the small intestine and colon. Immunofluorescence localized the hephaestin protein to the mature villus enterocytes with little or no expression in the crypts. Variations in iron status had a small but nonsignificant effect on hephaestin expression in the duodenum. The high sequence conservation between rat and mouse hephaestin is consistent with this protein playing a central role in intestinal iron absorption, although its precise function remains to be determined.

Amino Acid Sequence↗

Effect of mechanical deformation of neutrophils on their CD18/ICAM-1-dependent adhesion.

Mechanical deformation of polymorphonuclear leukocytes (PMN) changes their expression of the surface adhesion molecule CD11b/CD18. We tested the hypothesis that mechanical deformation of PMN enhances their adhesiveness. Purified human PMN were deformed through either 5- or 3-microm polycarbonate membrane filters and allowed to adhere to 96-well plates coated with human recombinant intercellular adhesion molecule-1 (ICAM-1). Flow cytometric studies showed that deformation of PMN increased CD11b/CD18 expression (P < 0.01). PMN adhesion to ICAM-1-coated plates was dependent on the magnitude of cell deformation (5 microm, 63.8 +/- 8.1%, P < 0.04; 3 microm, 232.4 +/- 20.9%, P < 0.01). Priming of PMN (0.5 nM N-formyl-methionyl-leucyl-phenylalanine) before deformation (5 microm) increased PMN adhesion (63.8 +/- 8.1 vs. 105.3 +/- 16.4%; P < 0.04). Stimulation (5% zymosan-activated plasma) of PMN after deformation resulted in increased adhesion, and the degree of increase was dependent on the magnitude of PMN deformation (stimulation, 50.6 +/- 4%; 5-microm filtration and stimulation, 62.9 +/- 6.6%; 3-microm filtration and stimulation, 249.9 +/- 24.2%; P < 0.01). This study shows that mechanical deformation of PMN causes an increase in PMN adhesiveness to ICAM-1 that was enhanced by both priming of PMN before deformation and stimulation after cell deformation.

CD18 Antigens↗

Breeding System and pollination of selected plants endemic to Juan Fernandez Islands.

We conducted field studies on the Juan Fernández Islands flora on the breeding system of 25 endemic species from 17 families. We recorded data on flower features, pollen and ovule number, pollen/ovule ratio, pollen size, self-compatibility, floral visitors, and pollination. Flowers are mostly hermaphrodite, inconspicuous, small, and green. Six species are dioecious. Over 80% of the cosexual species are self compatible. However, many species are dichogamous (mostly protandrous); thus, even the self-compatible species may require pollen transfer. Selfing through geitonogamy seems to be the most common system, and several species express mixed breeding systems. Floral visitors are uncommon to rare, except for two hummingbird species (one native and one endemic) that visit five species we studied. In more than 300 h of observation of flowers over three field seasons, we detected only 23 native insect visits representing ten species (Diptera, Lepidoptera, and Coleoptera). One species each of an introduced ant and an introduced bee were also observed on some flowers, all near the single human settlement of San Juan Bautista. Wind directly moving pollen, or indirectly via shaking the flowers, is the most important pollen distribution mechanism. The majority of the wind-pollinated species bear some typical anemophilous features, but also others not characteristic of wind pollination, that presumably represent the condition of their biotically pollinated ancestors. Floral features often reflect ancestral reproductive systems, so floral biology studies of oceanic islands in particular must be done with cognizance of presumed ancestral forms, because the observed characters can be misleading.

Journal Article↗

Jagged1 (JAG1) mutation detection in an Australian Alagille syndrome population.

Alagille syndrome (AGS) is an autosomal dominant disorder characterized by abnormal development of the liver, heart, skeleton, eye, and face. Mutations in the Jagged1 gene (JAG1) have been found to result in the AGS phenotype and both protein truncating mutations and missense mutations have been identified. Using single stranded conformational polymorphism analysis we have screened 22 AGS affected individuals from 19 families for mutations within Jagged1. Twelve distinct Jagged1 mutations were identified in 15 (68.2%) of the 22 AGS cases, seven of which are novel. The mutations include three small deletions (25%), two small insertions (16.6%), three missense mutations (25%), two nonsense mutations (16.6%), and two splice-site mutations (16.6%). These mutations are spread across the entire coding sequence of the gene and most are localized to highly conserved motifs of the protein predicted to be important for Jagged1 function. One-half of the mutations found in this study are located between exons 9 and 12, a region constituting only 12% of the coding sequence. A splice-donor site mutation in intron 11 was shown to cause aberrant splicing of Jagged1 mRNA, consequently terminating translation prematurely in exon 12. The results of this study are consistent with the proposal that either haploinsufficiency for wild type Jagged1 and/or dominant negative effects produced by mutated Jagged1 are responsible for the AGS phenotype.

Alagille Syndrome↗

Cheek cell phospholipids in human infants: a marker of docosahexaenoic and arachidonic acids in the diet, plasma, and red blood cells.

BACKGROUND: Assessment of essential fatty acid status requires collection of blood or adipose tissue samples. However, these invasive techniques cannot always be used in studies involving infants, young children, or subjects from whom it is difficult to obtain blood. A body tissue that is easy to access is the buccal mucosa (cheek cells). OBJECTIVE: The objective was to investigate the degree to which fatty acids of cheek cells reflect the fatty acid content of plasma, red blood cells, and the diet. DESIGN: Thirty-one infants aged 12 mo were enrolled. Five infants were fed human milk and 26 infants received formulas that provided a wide range of arachidonic acid and docosahexaenoic acid (DHA) intakes. Cheek cells were collected on a small piece of gauze by gently swabbing the inside of the cheek 3 times. Lipids were extracted from the gauze and the phospholipid fatty acid content of the cheek cells was determined. RESULTS: Cheek cell DHA and arachidonic acid in phospholipids were significantly correlated with DHA and arachidonic acid in plasma [r = 0.61 (P < 0.001) and r = 0.37 (P <0.05), respectively], red blood cells [r = 0.58 (P < 0.001) and r = 0.37 (P < 0.05), respectively], and the diet [r = 0.65 (P < 0.001) and r = 0. 51 (P < 0.01), respectively]. CONCLUSIONS: Given these correlations and the ease and noninvasive nature of this technique, cheek cell fatty acids may serve as a marker of the essential fatty acid content, especially of DHA and arachidonic acid, in plasma, red blood cells, and the diet.

Analysis of Variance↗

Characterization of the extended-spectrum beta-lactamase reference strain, Klebsiella pneumoniae K6 (ATCC 700603), which produces the novel enzyme SHV-18.

Klebsiella pneumoniae K6 (ATCC 700603), a clinical isolate, is resistant to ceftazidime and other oxyimino-beta-lactams. A consistent reduction in the MICs of oxyimino-beta-lactams by at least 3 twofold dilutions in the presence of clavulanic acid confirmed the utility of K. pneumoniae K6 as a quality control strain for extended-spectrum beta-lactamase (ESBL) detection. Isoelectric-focusing analysis of crude lysates of K6 demonstrated a single beta-lactamase with a pI of 7.8 and a substrate profile showing preferential hydrolysis of cefotaxime compared to ceftazidime. PCR analysis of total bacterial DNA from K6 identified the presence of a bla(SHV) gene. K6 contained two large plasmids with molecular sizes of approximately 160 and 80 kb. Hybridization of plasmid DNA with a bla(SHV)-specific probe indicated that a bla(SHV) gene was encoded on the 80-kb plasmid, which was shown to transfer resistance to ceftazidime in conjugal mating experiments with Escherichia coli HB101. DNA sequencing of this bla(SHV)-related gene revealed that it differs from bla(SHV-1) at nine nucleotides, five of which resulted in amino acid substitutions: Ile to Phe at position 8, Arg to Ser at position 43, Gly to Ala at position 238, and Glu to Lys at position 240. In addition to the production of this novel ESBL, designated SHV-18, analysis of the outer membrane proteins of K6 revealed the loss of the OmpK35 and OmpK37 porins.

Amino Acid Sequence↗

Quantitative description of the far-lateral transcondylar transtubercular approach to the foramen magnum and clivus.

OBJECT: The purpose of this study was to evaluate the far-lateral transcondylar transtubercular approach (complete FLA) based on quantitative measurements of the exposure of the foramen magnum and petroclival area obtained after each successive step of this approach. METHODS: The complete FLA was reproduced in eight specially prepared cadaveric heads (a total of 15 sides). The approach was divided into six steps: 1) C-1 hemilaminectomy and suboccipital craniectomy with unroofing of the sigmoid sinus (basic FLA); 2) partial resection of the occipital condyle (up to the hypoglossal canal); 3) removal of the jugular tuberculum; 4) mastoidectomy (limited to the labyrinth and the fallopian canal) and retraction of the sigmoid sinus; 5) resection of the lateral mass of C-1 with mobilization of the vertebral artery; and 6) resection of the remaining portion of the occipital condyle. After each successive step, a standard set of measurements was obtained using a frameless stereotactic device. The measurements were used to estimate two parameters: the size of the exposed petroclival area and the size of a spatial cone directed toward the anterior rim of the foramen magnum, which depicts the amount of surgical freedom available for manipulation of instruments. The initial basic FLA provided exposure of only 21 +/- 6% of the petroclival area that was exposed with the full, six-step maximally aggressive (complete) FLA. Likewise, only 18 +/- 9% of the final surgical freedom was obtained after the basic FLA was performed. Each subsequent step of the approach increased both petroclival exposure and surgical freedom. The most dramatic increase in petroclival exposure was noted after removal of the jugular tuberculum (71 +/- 12% of final exposure), whereas the least improvement in exposure occurred after the final step, which consisted of total condyle resection. CONCLUSIONS: The complete FLA provides wide and sufficient exposure of the foramen magnum and lower to middle clivus. The complete FLA consists of several steps, each of which contributes to increasing petroclival exposure and surgical freedom. However, the FLA may be limited to the less aggressive steps, while still achieving significant exposure and surgical freedom. The choice of complete or basic FLA thus depends on the underlying pathological condition and the degree of exposure required for effective surgical treatment.

Algorithms↗

Classification and quantification of the petrosal approach to the petroclival region.

OBJECT: The petrosal approach to the petroclival region has been used by a variety of authors in various ways and the terminology has become quite confusing. A systematic assessment of the benefits and limitations of each approach is also lacking. The authors classify their approach to the middle and upper clivus, review the applications for each, and test their hypotheses on a cadaver model by using frameless stereotactic guidance. METHODS: The petrosal approach to the upper and middle clivus is divided into four increasingly morbidity-producing steps: retrolabyrinthine, transcrusal (partial labyrinthectomy), transotic, and transcochlear approaches. Four latex-injected cadaveric heads (eight sides) underwent dissection in which frameless stereotactic guidance was used. An area of exposure 10 cm superficial to a central target (working area) was calculated. The area and length of clival exposure with each subsequent dissection was also calculated. The retrolabyrinthine approach spares hearing and facial function but provides for only a small window of upper clival exposure. The view afforded by what we have called the transcrusal approach provides for up to four times this exposure. The transotic and transcochlear procedures, although producing more morbidity, add little in terms of a larger clival window. However, with each step, the surgical freedom for manipulation of instruments increases. CONCLUSIONS: The petrosal approach to the upper and middle clivus is useful but should be used judiciously, because levels of morbidity can be high. The retrolabyrinthine approach has limited utility. For tumors without bone invasion, the transcrusal approach provides a much more versatile exposure with an excellent chance of hearing and facial nerve preservation. The transotic approach provides for greater versatility in treating lesions but clival exposure is not greatly enhanced. Transcochlear exposure adds little in terms of intradural exposure and should be reserved for cases in which access to the petrous carotid artery is necessary.

Cranial Fossa, Posterior↗

RAPD marker diversity within and divergence among species of Dendroseris (Asteraceae: Lactuceae).

Random Amplified Polymorphic DNA (RAPD) markers were used to measure genetic diversity within and divergence among species of Dendroseris (Asteraceae: Lactuceae), a genus endemic to the Juan Fernandez Islands, Chile. Results were compared to previous studies employing allozymes. For five of the species, RAPD banding patterns distinguished all individuals examined, and different mutilocus genotypes were found even in species exhibiting no allozyme diversity. RAPD band diversities ranged from 0.003 to 0.022 within species; >90% of total diversity was among species and <10% within them. Relative levels of allozyme and RAPD diversity were similar for some species, particularly those with highest and lowest diversities, but overall there was no significant correlation. Relationships inferred from a neighbor-joining tree generated from RAPD bands were similar to results obtained from morphology, chloroplast DNA (cpDNA) restriction site mutations, and sequences from the internal transcribed spacer regions of nuclear ribosomal DNA (ITS), but somewhat better resolution was achieved. Relationships shown by allozymes differed from trees based on other data; this ostensibly is a result of the sharing of ancestral alleles and the absence of alleles generated subsequent to speciation. Dendroseris represents an example where RAPD markers, because of their greater variability, provide a useful alternative to allozymes for assessing diversity in rare species endemic to oceanic islands and for resolving relationships among the species.

Journal Article↗

Hephaestin, a ceruloplasmin homologue implicated in intestinal iron transport, is defective in the sla mouse.

Iron is essential for many cellular functions; consequently, disturbances of iron homeostasis, leading to either iron deficiency or iron overload, can have significant clinical consequences. Despite the clinical prevalence of these disorders, the mechanism by which dietary iron is absorbed into the body is poorly understood. We have identified a key component in intestinal iron transport by study of the sex-linked anaemia (sla) mouse, which has a block in intestinal iron transport. Mice carrying the sla mutation develop moderate to severe microcytic hypochromic anaemia. Although these mice take up iron from the intestinal lumen into mature epithelial cells normally, the subsequent exit of iron into the circulation is diminished. As a result, iron accumulates in enterocytes and is lost during turnover of the intestinal epithelium. Biochemical studies have failed to identify the underlying difference between sla and normal mice, therefore, we used a genetic approach to identify the gene mutant in sla mice. We describe here a novel gene, Heph, encoding a transmembrane-bound ceruloplasmin homologue that is mutant in the sla mouse and highly expressed in intestine. We suggest that the hephaestin protein is a multicopper ferroxidase necessary for iron egress from intestinal enterocytes into the circulation and that it is an important link between copper and iron metabolism in mammals.

Amino Acid Sequence↗

Genetics of hemochromatosis.

Hereditary hemochromatosis (HHC) is a common autosomal recessive disorder of iron metabolism that results in progressive iron overload and can be fatal if untreated. The hemochromatosis gene (HFE) was identified by positional cloning in 1996. Two missense mutations have been described in HFE. The majority of HHC patients are homozygous for a cysteine-to-tyrosine substitution (C282Y); however, a small number are homozygous for a histidine-to-aspartic-acid substitution (H63D) or are heterozygous for both of these mutations. Mechanisms by which C282Y and H63D may disrupt the normal functioning of HFE have been suggested, but the role of HFE in the process of normal iron metabolism has yet to be clearly defined.

Amino Acid Substitution↗

Comparison of three techniques for calculation of target coordinates in functional stereotactic procedures.

We compared three techniques of target coordinate determination for various functional stereotactic procedures. All procedures were based on preoperative MRI with contiguous 3-mm cuts. The first technique involved determination of anatomical landmarks and fiducial markers of the stereotactic frame on the monitor screen of an MRI scanner and calculation of the target point using a series of formulas; the second technique used a Leksell tabletop localizer, and the third technique is a part of 'Stealth' stereotactic navigation software. Final coordinates for the procedure were derived from all three techniques and subsequently adjusted using intraoperative electrical macrostimulation. We found that difference between techniques was on average 0.9 +/- 0.4 mm in each of three directions, and 1.8 +/- 0.9 mm in absolute distance. There were 7 cases in which one of the techniques had a discrepancy of more than 3 mm (more than 1 MRI slice thickness) compared with the other two, indicating a potential error in coordinate determination. This difference could potentially result in inappropriate placement of the electrode, thus affecting the procedure outcome. In 6 cases, such an error apparently occurred with the first or second technique of calculation. The average number of mapping trajectories decreased from 1.8 to 1.4 since this stereotactic software became a part of operative planning. We conclude that use of computerized planning software increases the precision of target coordinate calculation and improves the accuracy of functional stereotactic procedures.

Analog-Digital Conversion↗

Quantification of increased exposure resulting from orbital rim and orbitozygomatic osteotomy via the frontotemporal transsylvian approach.

OBJECT: Use of orbital rim and orbitozygomatic osteotomy has been extensively reported to increase exposure in neurosurgical procedures. However, there have been few attempts to quantify the extent of additional exposure gained by these maneuvers. Using a novel laboratory technique, the authors have attempted to measure the increase in the "area of exposure" that is gained by removal of the orbital rim and zygomatic arch via the frontotemporal transsylvian approach. METHODS: The authors dissected five cadavers bilaterally. The area of exposure provided by the frontotemporal transsylvian approach was determined by using a frameless stereotactic device. With the tip of a microdissector placed on targets deep within the exposure, the position of the end of the microdissector handle was measured in three-dimensional space as the microdissector was rotated around the periphery of the operative field. This maneuver was performed via the frontotemporal approach alone as well as with orbital rim and orbitozygomatic osteotomy approaches. After data manipulation, the areas of exposure corresponding to the polygons used to define these handle positions were calculated and directly compared. On average, the area of exposure provided by the frontotemporal transsylvian approach was increased 26 to 39% (p<0.05) by adding orbital rim osteotomy and an additional 13 to 22% (not significant) with removal of the zygomatic arch. CONCLUSIONS: Significant and consistent increases in surgical exposure were obtained by using orbital osteotomy, whereas zygomatic arch removal produced less consistent gains. Both maneuvers may be expected to improve surgical access. However, because larger and more consistent gains were afforded by orbital rim removal, the threshold for removal of this portion of the orbitozygomatic complex should be lower.

Brain Mapping↗

Reproductive biology of Lactoris fernandeziana (Lactoridaceae).

Lactoris fernandeziana, monotypic in its family, is endemic to the cloud forests of Robinson Crusoe Island. Although there has been considerable study of the relationships of Lactoris, as a rare species and as a putative primitive paleoherb, little is known of its reproductive biology. Knowledge of the latter is essential for effective conservation programs. The species is gynomonoecious. The overall proportion of flowers is ∼1 female:1 hermaphrodite. The inconspicuous semipendulous green flowers, usually in mixed-gender inflorescences, do not produce rewards. Hermaphrodite flowers are herkogamous and protogynous. Pollen grains are shed from the extrorse anthers in permanent dry tetrads. There is a mean of 12879 tetrads per hermaphrodite flower. Both flower types bear an average of ∼18 ovules. The P/O (pollen/ovule) ratios imply facultative or obligate xenogamy, but hand pollinations show that Lactoris is self-compatible. No floral visitors were ever observed, but stigmata of open-pollinated flowers bore tetrads, and 64% of such styles had pollen tubes. Flowers enclosed in large mesh (1 mm) bags bore similar numbers of tetrads and pollen tubes. Thus, we conclude that Lactoris is anemophilous, a syndrome perhaps reflected by the P/O ratio. Low genetic diversity (isozymes and DNA) supports selfing and implies limited distance wind pollen dispersal. The small size of the island, the ± 1000 extant Lactoris plants, coupled with anemophily, self-compatibility, and pendant flower position, have yielded a geitonogamous system with high seed set and low genetic diversity. If inbreeding depression is expressed, it is in seed germination and seedling vigor, for Lactoris is very difficult to cultivate. For this species, effective conservation practices need to focus on habitat preservation and promotion of outcrossing.

Journal Article↗