Correction: Therapeutic targeting of Id2 reduces growth of human colorectal carcinoma in the murine liver.

An amendment to this paper has been published and can be accessed via a link at the top of the paper.

Ranavirus: past, present and future.

Emerging infectious diseases are a significant threat to global biodiversity. While historically overlooked, a group of iridoviruses in the genus Ranavirus has been responsible for die-offs in captive and wild amphibian, reptile and fish populations around the globe over the past two decades. In order to share contemporary information on ranaviruses and identify critical research directions, the First International Symposium on Ranaviruses was held in July 2011 in Minneapolis, MN, USA. Twenty-three scientists and veterinarians from nine countries examined the ecology and evolution of ranavirus-host interactions, potential reservoirs, transmission dynamics, as well as immunological and histopathological responses to infection. In addition, speakers discussed possible mechanisms for die-offs, and conservation strategies to control outbreaks.

Therapeutic targeting of Id2 reduces growth of human colorectal carcinoma in the murine liver.

During development inhibitor of DNA-bind-2 (Id2) regulates proliferation and differentiation. Id2 expression has been detected in cancer cells, yet its cellular function and validity as a therapeutic target remains largely unknown. Immunohistochemical analysis of colorectal cancer (CRC) specimens revealed that Id2 was undetectable in normal colonic mucosa, but occurs in 40% of primary tumors and in most CRC liver metastases (P<0.0001). Additionally, Id2 was expressed in all CRC cell lines assayed. CRC cells with reduced Id2 expression demonstrated reduced proliferation. Analysis of CRC cell cycle regulatory proteins showed that reducing Id2 levels reduces cyclin D1 levels and increased p21 levels. Reduction of Id2 expression also enhanced tumor cell apoptosis, increasing levels of the pro-apoptotic protein Bim/Bod, and cleavage of caspase-7 and poly (ADP-ribose) polymerase. In vivo studies show tumors derived from cells with decreased Id2 levels formed smaller tumors with fewer metastases compared with tumors with normal levels (P<0.05). Furthermore, intraperitoneal administration of Id2 small interfering RNA (siRNA) conjugated with the neutral liposome 1,2-dioleoyl-sn-glycero-3-phosphatidylcholine decreased tumor burden in mice compared with control treatment (P=0.006). We conclude that Id2 is upregulated in CRC, and is important in promoting cell survival. In vivo targeting of Id2 by siRNA establishes that it is a valid therapeutic target where its expression occurs.

The determination of glycyrrhizic acid in Glycyrrhiza uralensis Fisch. ex DC. (Zhi Gan Cao) root and the dried aqueous extract by LC-DAD.

A rapid, sensitive and specific reversed phase high-performance liquid chromatographic (LC) method with photodiode array detection (DAD) has been developed for the determination of glycyrrhizic acid in both the raw herb and a commercially prepared dried aqueous extract of Glycyrrhiza uralensis Fisch. ex DC. root (Zhi Gan Cao, liquorice). It was determined that extracting the raw herb in aqueous methanol (50:50 v/v) by sonication for 2 x 30 min was the most efficient sample preparation. Baseline resolution of the glycyrrhizic acid peak was achieved on a Varian Polaris RP C18-A (250 mm x 4.6 mm, 5 microm packing) column using an isocratic mobile phase consisting of 0. v aqueous phosphoric acid and acetonitrile in the ratio 60:40 v/v. Chromatograms were monitored between 200 and 400 nm for peak purity assessments, with quantitation performed at 254 nm. Glycyrrhizic acid calibration curves in the concentration range of 14-558 microg/ml were prepared on the day of analysis. Curve fitting was by the least-squares method, with correlation coefficients of >0.9998 obtained each time. The average recovery at three spike levels (50, 100, 200%) was of 95.91+/-1.05% and 98.36+/-3.45% (+/-S.D., n=7) for the spiked raw herb and dried aqueous extract respectively. The limit of detection and limit of quantitation was 0.52 and 1.72 mg/g respectively for the raw herb, and 0.75 and 2.51 mg/g respectively for the dried aqueous extract. Identity confirmation of the chromatographic peak was achieved by (-) electrospray ionisation tandem mass spectrometry. The concentration of glycyrrhizic acid in the root and dried aqueous extract was found to be 31.1+/-0.2 and 40.4+/-0.3mg/g (+/-S.D., n=7) respectively.

Genomic cartography of varicella-zoster virus: a complete genome-based analysis of strain variability with implications for attenuation and phenotypic differences.

In order to gain a better perspective on the true variability of varicella-zoster virus (VZV) and to catalogue the location and number of differences, 11 new complete genome sequences were compared with those previously in the public domain (18 complete genomes in total). Three of the newly sequenced genomes were derived from a single strain in order to assess variations that can occur during serial passage in cell culture. The analysis revealed that while VZV is relatively stable genetically it does posses a certain degree of variability. The reiteration regions, origins of replication and intergenic homopolymer regions were all found to be variable between strains as well as within a given strain. In addition, the terminal viral sequences were found to vary within and between strains specifically at the 3' end of the genome. Analysis of single nucleotide polymorphisms (SNPs) identified a total of 557 variable sites, 451 of which were found in coding regions and resulted in 187 different in amino acid substitutions. A comparison of the SNPs present in the two gE mutant strains, VZV-MSP and VZV-BC, suggested that the missense mutation in gE was primarily responsible for the accelerated cell spread phenotype. Some of the variations noted with high passage in cell culture are consistent with variations seen in the IE62 gene of the vaccine strains (S628G, R958G and I1260V) that may help in pinpointing variations essential for attenuation. Although VZV has been considered to be one of the most genetically stable human herpesviruses, this initial assessment of genomic VZV cartography provides insight into ORFs with previously unreported variations.

A column capacity study of single, serial, and parallel linked rod monolithic high performance liquid chromatography columns.

The loading capacity of rod monolithic C18 columns was found to be sensitive to the injection volume, but essentially insensitive to the mass loading for a separation of oligostyrenes. When rod monoliths were coupled in series the injection volume loading increased, as too did the resolution of the oligomers, but at the expense of separation time. The volume load capacity of these serially connected monoliths was, however, not directly proportional to the number of columns connected. The volume load capacity was, however, directly proportional to the number of columns when the monoliths were connected in parallel and the flow stream split between each of the monolithic channels. When the number of monoliths in each channel equaled the number of monoliths that were connected in a single channel serial system the peak capacity and retention time was equivalent for both systems, but the volume load capacity of the parallel system was twice that of the serial connection each time the number of channels doubled. The results of this study indicate that parallel connection of rod monolithic columns may be useful for preparative scale and multidimensional separations where it is important that the volume load capacity is high.

Chronic axillary wound repair in a cat with omentalisation and omocervical skin flap.

A mature, neutered male domestic cat was presented with a chronic axillary wound of unknown origin and at least three years' duration. The diagnostic investigation included screening tests for feline leukaemia virus and feline immunodeficiency virus, tissue culture and histological examination. No underlying aetiology or perpetuating cause could be identified. An omental flap was created, passed via a subcutaneous tunnel, and packed into the wound site after excision of all chronic granulation tissue. The skin defect was closed using an omocervical axial pattern skin flap. A small area of the distal edge of the flap became necrotic but the defect healed by second intention. To the author's knowledge, this is the first clinical report of the use of an omocervical skin flap for repair of a chronic axillary wound in a cat. This flap offers a useful alternative where the use of a thoracodorsal axial pattern flap is not possible due to the extent of the lesion.

Overexpression of neuropilin-1 promotes constitutive MAPK signalling and chemoresistance in pancreatic cancer cells.

Neuropilin-1 (NRP-1) is a novel co-receptor for vascular endothelial growth factor (VEGF). Neuropilin-1 is expressed in pancreatic cancer, but not in nonmalignant pancreatic tissue. We hypothesised that NRP-1 expression by pancreatic cancer cells contributes to the malignant phenotype. To determine the role of NRP-1 in pancreatic cancer, NRP-1 was stably transfected into the human pancreatic cancer cell line FG. Signal transduction was assessed by Western blot analysis. Susceptibility to anoikis (detachment induced apoptosis) was evaluated by colony formation after growth in suspension. Chemosensitivity to gemcitabine or 5-fluorouracil (5-FU) was assessed by MTT assay in pancreatic cancer cells following NRP-1 overexpression or siRNA-induced downregulation of NRP-1. Differential expression of apoptosis-related genes was determined by gene array and further evaluated by Western blot analysis. Neuropilin-1 overexpression increased constitutive mitogen activated protein kinase (MAPK) signalling, possibly via an autocrine loop. Neuropilin-1 overexpression in FG cells enhanced anoikis resistance and increased survival of cells by > 30% after exposure to clinically relevant levels of gemcitabine and 5-FU. In contrast, downregulation of NRP-1 expression in Panc-1 cells markedly increased chemosensitivity, inducing > 50% more cell death at clinically relevant concentrations of gemcitabine. Neuropilin-1 overexpression also increased expression of the antiapoptotic regulator, MCL-1. Neuropilin-1 overexpression in pancreatic cancer cell lines is associated with (a) increased constitutive MAPK signalling, (b) inhibition of anoikis, and (c) chemoresistance. Targeting NRP-1 in pancreatic cancer cells may downregulate survival signalling pathways and increase sensitivity to chemotherapy.

A biomechanical investigation of the static stabilisers of the glenohumeral joint in the dog.

The aim was to determine the relative contribution of the glenohumeral ligaments/joint capsule complex and the limited joint volume/adhesion-cohesion (UV/AC) mechanisms to the static stability of the canine glenohumeral joint (GHJ). The GHJ of 32 complete cadaver specimens were manipulated to determine the presence of gross laxity (> 2mm translation) at joint angles of 150 degrees, 135 degrees and 90 degrees. Following the removal of the peri-articular muscles, laxity was measured by applying a 15N force to the scapula in a variety of directions whilst the humerus was fixed to a jig. This was repeated for the abovementioned angles in intact joints and after venting and flushing to eliminate the UV/AC mechanisms. Results of clinical palpation and biomechanical studies were compared. Manipulation revealed a marked variation between joints although there was a tendency for progressive laxity as the joint was flexed. This finding was supported by the biomechanical study that also demonstrated no significant difference between intact and vented joints except in a cranio-caudal direction at 135 degrees and 90 degrees, and in a medial/lateral direction at 135 degrees. Static stabilisers permitted a wide range of motion. Individual clinical palpations did not correlate well with biomechanical findings. Manipulation was difficult to perform and is unlikely to be a reliable method for determining subtle joint laxity. The static stabilisers do not appear to play a significant role in providing joint constraint during normal range of motion (except extension). Diagnoses of pathological laxity attributable to the static stabilisers should be made with caution and surgical treatments should take cognizance of the above-mentioned biomechanical features.

Metacercariae of Clinostomum attenuatum in Ambystoma tigrinum mavortium, Bufo cognatus and Spea multiplicata from west Texas.

Tissues from barred tiger salamanders (Ambystoma tigrinum mavortium), Great Plains toads (Bufo cognatus) and New Mexico spadefoots (Spea multiplicata) collected from 16 playa wetlands in Texas during 1999 and 2000 were examined by light microscopy. Digenean cysts were primarily distributed subcutaneously throughout the specimens and occasionally coelomic invasion was noted. The parasites within the cysts were 1.5-2 mm in diameter, with a thin (c. 10 microm wide) eosinophilic-staining tegument, two suckers (oral and ventral), posteriorly located primordial genitalia and paired digestive caeca. These digeneans were identified as the metacercariae of Clinostomum attenuatum. This is the first record of Clinostomum attenuatum in these amphibian species.

Cervical spondylomyelopathy (wobbler syndrome) in the Boerboel.

The Boerboel is a South African large-breed dog resembling a Bullmastiff. The records of Onderstepoort Veterinary Academic Hospital were searched for dogs that had presented, between 1998 and 2003, with symptoms indicative of wobbler syndrome and had undergone survey radiographic and myelographic studies. Ten cases fitted the inclusion criteria. Dogs presented within the first 2 years of life, often with acute onset of symptoms. All presented with pelvic limb and 6 with concomitant thoracic limb ataxia or paresis. Treatment varied and included none (4), prednisolone (2), and dorsal laminectomy (2). Two dogs were euthanased at the time of diagnosis. The breed appears to be affected with a form of spondylomyelopathy that comprises bony malformation of cervical and/or thoracic vertebrae. In 8 dogs, malformations were evident on survey radiographs and were characterised by enlarged, irregular articular facets and associated medial deviation of the pedicles. These changes resulted in axial compression of the spinal cord best seen on ventrodorsal or dorsoventral myelographic studies. Multiple vertebrae were affected in some dogs and lesions were not confined to the caudal area of the cervical spine. Three dogs were alive and without symptoms at follow-up. Four were euthanased as a result of the disease and 1 died as a result of post-operative complications. Two additional dogs presenting with wobbler clinical signs are also described. One had medial deviation of the T5 and T6 caudal pedicles and 1 dog suffered from multiple cervical articular facet synovial cysts.

Characterization of chocolate milk spoilage patterns.

Standard plate counts (SPC) and psychrotrophic plate counts (PPC) from chocolate milk samples were compared with those of unflavored milk samples plated within 24 h of processing and at 7, 10, and 14 days of storage at 6 degrees C using matched samples collected over four time periods from four milk-processing plants. Bacterial numbers within 24 h of processing were not significantly different in unflavored and in chocolate milk samples (P > 0.001), with SPC less than 1,000 CFU/ml and PPC below 10 CFU/ml for both types of products. SPC and PPC were higher in chocolate milk samples than in unflavored milk samples collected from all four plants after 14 days of storage (P < 0.001). To examine the effects of chocolate milk components on bacterial numbers, SPC for days 0, 7, 14, and 21 were monitored in samples of experimentally prepared unflavored milk, milk with chocolate powder and sucrose (chocolate milk), milk with sucrose only, and milk containing chocolate powder only. At days 14 and 21, SPC were higher in both chocolate milk and in milk with chocolate powder only, than in either the unflavored milk or milk with sucrose only (P < 0.001). These findings suggest that the addition of chocolate powder to milk can contribute to a greater relative increase in bacterial numbers in pasteurized chocolate milk than in identically processed unflavored milk at 14 days postprocessing.

Blood parameters of clinically normal captive bobcats (Felis rufus).

Data on blood parameters were obtained from 56 (32 female, 24 male) clinically healthy, captive bobcats (Felis rufus). The animals were housed under ambient conditions at the Mississippi State University Captive Bobcat Facility. Samples were collected opportunistically between 1982 and 1995. Differences (P < or = 0.05) in blood parameters existed between sexes, among age classes (kitten, subadult, adult, and geriatric), and among stages in the reproductive cycle (breeding, parturition, recycling, and anestrus). Blood parameters for bobcats were similar to reported values for domestic cats (Felis catus). These results can be used as as reference to indicate when blood parameter aberrations exist in bobcats.

Early environmental influences can attenuate the blood pressure response to acute stress in borderline hypertensive rats.

The objective of this experiment was to test the hypothesis that early environmental influences could affect cardiovascular reactivity to acute stress in borderline hypertensive rats. This study utilized a cross-fostering paradigm in which borderline hypertensive rat (BHR) pups were reared by their natural spontaneously hypertensive rat (SHR) mothers, by an adoptive SHR dam, or by an adoptive Wistar-Kyoto female. Several maternal behavior measures were taken throughout weaning. Cardiovascular responses to acute stress were measured in adult BHR offspring. SHR dams spent significantly more time engaged in arched, as contrasted with passive, nursing behaviors throughout the preweaning period. Although strain of dam had no effect on resting blood pressure or heart rate, BHR reared by WKY dams had a greatly reduced blood pressure response to acute stress compared with BHR raised by either natural or adoptive SHR dams. These results suggest that the expression of cardiovascular characteristics in the BHR can be modified by early environmental influences and support the hypothesis that the expression of the hypertensive phenotype in the BHR is dependent upon information that is conveyed both genetically and environmentally.

Effect of the menstrual cycle on detection and typing of human papillomavirus in uterine cervical cells.

We observe fluctuations in human papillomavirus detection and variation in genotyping between sequential cervical cell specimens analyzed by filter in situ hybridization. Furthermore, specimen adequacy for analysis varies. To determine whether these phenomena are correlated with menstrual cycle stage at the time of sampling, we analyzed cervical cell specimens from women with cervical intraepithelial neoplasia. Specimens were categorized on the basis of a 28-day menstrual cycle and were analyzed by hybridization to combined probes for virus types 6 and 11 or types 16 and 18. Specimen adequacy was determined by hybridization to a human Alu I repetitive deoxyribonucleic acid probe. Analysis of data with chi 2 revealed that fluctuations in virus detection and type variation are unrelated to menstrual cycle stage. Specimen adequacy is stage-dependent for women who take oral contraceptives. Whereas specimens can be collected at any time other than the first week of the menstrual cycle, accurate determination of infection status requires multiple assessments.