In vivo growth and genomic characterization of rickettsia-like organisms isolated from farmed Chinook salmon (Oncorhynchus tshawytscha) in New Zealand.

A rickettsia-like organism, designated NZ-RLO2, was isolated from Chinook salmon (Oncorhynchus tshawytscha) farmed in the South Island, New Zealand. In vivo growth showed NZ-RLO2 was able to grow in CHSE-214, EPC, BHK-21, C6/36 and Sf21 cell lines, while Piscirickettsia salmonis LF-89T grew in all but BHK-21 and Sf21. NZ-RLO2 grew optimally in EPC at 15°C, CHSE-214 and EPC at 18°C. The growth of LF-89 T was optimal at 15°C, 18°C and 22°C in CHSE-24, but appeared less efficient in EPC cells at all temperatures. Pan-genome comparison of predicted proteomes shows that available Chilean strains of P. salmonis grouped into two clusters (p-value = 94%). NZ-RLO2 was genetically different from previously described NZ-RLO1, and both strains grouped separately from the Chilean strains in one of the two clusters (p-value = 88%), but were closely related to each other. TaqMan and Sybr Green real-time PCR targeting RNA polymerase (rpoB) and DNA primase (dnaG), respectively, were developed to detect NZ-RLO2. This study indicates that the New Zealand strains showed a closer genetic relationship to one of the Chilean P. salmonis clusters; however, more Piscirickettsia genomes from wider geographical regions and diverse hosts are needed to better understand the classification within this genus.

Congenital-infantile fibrosarcoma of the foot--avoidance of amputation.

Congenital-infantile fibrosarcoma is a rare entity with a five year survival rate of over 90%. Surgery is still the most common treatment modality with amputation often necessary. There have been reports supporting the use of neoadjuvant chemotherapy to debulk the tumour in an effort to facilitate limb sparing surgery. We report a case of a newborn who presented with a life threatening haemorrhage from a fibrosarcoma of the foot, successfully treated with Vincristine, Actinomycin and Cyclophosphamide (VAC) chemotherapy alone.

A multicenter, randomized phase 2 study to establish combinations of CBP501, cisplatin and nivolumab for ≥3rd-line treatment of patients with advanced pancreatic adenocarcinoma.

BACKGROUND: There is no standard of care for ≥ 3rd-line treatment of metastatic pancreatic adenocarcinoma (PDAC). CBP501 is a novel calmodulin-binding peptide that has been shown to enhance the influx of platinum agents into tumor cells and tumor immunogenicity. This study aimed to (1) confirm efficacy of CBP501/cisplatin/nivolumab for metastatic PDAC observed in a previous phase 1 study, (2) identify combinations that yield 35% 3-month progression-free survival rate (3MPFS) and (3) define the contribution of CBP501 to the effects of combination therapy. METHODS: CBP501 16 or 25 mg/m2 (CBP(16) or CBP(25)) was combined with 60 mg/m2 cisplatin (CDDP) and 240 mg nivolumab (nivo), administered at 3-week intervals. Patients were randomized 1:1:1:1 to (1) CBP(25)/CDDP/nivo, (2) CBP(16)/CDDP/nivo, (3) CBP(25)/CDDP and (4) CDDP/nivo, with randomization stratified by ECOG PS and liver metastases. A Fleming two-stage design was used, yielding a one-sided type I error rate of 2.5% and 80% power when the true 3MPFS is 35%. RESULTS: Among 36 patients, 3MPFS was 44.4% in arms 1 and 2, 11.1% in arm 3% and 33.3% in arm 4. Two patients achieved a partial response in arm 1 (ORR 22.2%; none in other arms). Median PFS and OS were 2.4, 2.1, 1.5 and 1.5 months and 6.3, 5.3, 3.7 and 4.9 months, respectively. Overall, all treatment combinations were well tolerated. Most treatment-related adverse events were grade 1-2. CONCLUSIONS: The combination CBP(25)/(16)/CDDP/nivo demonstrated promising signs of efficacy and a manageable safety profile for the treatment of advanced PDAC. CLINICAL TRIAL REGISTRATION: NCT04953962.

Culture conditions affect the chemical composition of the exopolysaccharide synthesized by the fungus Aureobasidium pullulans.

AIMS: To identify if culture conditions affect the chemical composition of exopolysaccharide (EPS) produced by Aureobasidium pullulans. METHODS AND RESULTS: In batch airlift and continuously stirred tank (CSTR) reactors the EPS produced with low (0.13 g l(-1) N) initial NaNO(3) or (NH(4))(2)SO(4) levels contained pullulan, with maltotriose as its major component, similar to that synthesized in the airlift reactor with high (0.78 g l(-1) N) initial NaNO(3) levels. EPS produced by CSTR grown cultures with high (NH(4))(2)SO(4) levels contained little pullulan, possibly because of a population shift from unicells to mycelium. This chemical difference may explain why total EPS yields did not fall as they did with cultures grown under identical conditions with high NaNO(3) levels, where the pullulan component of the EPS disappeared. EPS synthesized in N-limiting chemostat cultures of A. pullulans changed little with growth rate or N source, being predominantly pullulan consisting of maltotriose units. CONCLUSIONS: While the EPS chemical composition changed little under N-limiting conditions, high initial medium N levels determined maltotriose content and/or pullulan content possibly by dictating culture morphology. SIGNIFICANCE AND IMPACT OF THE STUDY: These results emphasize the requirement of all studies to determine EPS chemical composition when examining the influence of culture conditions on EPS yields.

Occupational injuries in foreign-national workers presenting to St James's Hospital Plastic Surgery service.

This study was undertaken to investigate our impression that migrant foreign-national workers were more at risk of sustaining work place injuries requiring referral to our Plastic Surgery service than their indigenous Irish counterparts. Data were collected prospectively from August 2006 to February 2007 on all work-related injuries presenting to the Plastic Surgery service in St James's Hospital, Dublin. 201 work-related injuries were recorded during the six month study period. 40% (n = 81) of the study group were foreign-national workers. Foreign-national workers account for only nine percent of the total Irish workforce. 31% (n = 25) of the study group required a translator. Over half (55%) of all the foreign-national workers in the current study had been in their present job for less than six months at the time of injury compared to only nine percent of Irish workers. This study highlights that foreign-national workers in Ireland are at a disproportionately high risk of occupational injury when compared to their Irish colleagues and emphasises the need for targeted occupational health and safety measures in this vulnerable group.

Patterns of crystallin distribution in porcine eye lenses.

PURPOSE: To measure the protein distribution patterns in single young porcine lenses. METHODS: Twenty fresh porcine lenses from 5 to 6 months old animals were fractionated into 8-10 concentric fractions by controlled dissolution in phosphate buffer. Proportions of soluble and insoluble protein were determined by Bradford assay. Water-soluble proteins in all layers were separated into HMW, MMW, and LMW fractions by size-exclusion HPLC and constituents of each class further characterized by SDS gel electrophoresis, as were the water-insoluble proteins. Size-exclusion fractions were further separated by reverse-phase HPLC and the molecular masses of each peak determined by MALDI-TOF mass spectrometry. RESULTS: The major soluble proteins in the porcine lens are beta-crystallins. They comprise around 45% of the total protein in the outer lens decreasing gradually to 35% in the central region. Soluble alpha-crystallins vary from 35% to 22% from outer to inner lens. The proportion of soluble gamma-crystallin levels, substantially lower than that of the other protein classes, increases gradually with progression into the lens center. Insoluble protein levels also increase from outer to inner lens layers. CONCLUSIONS: In the young porcine lens, there is relative constancy in the levels of all three crystallin classes in the outer lens with alpha- and beta-crystallins representing the predominant protein classes. The increase in gamma-crystallin in the inner lens may contribute to the refractive index gradient.

Intraoperative retrograde mesenteric angioplasty for acute occlusive mesenteric ischaemia: a case series.

Acute mesenteric ischaemia secondary to atherosclerotic disease of the superior mesenteric artery is a surgical emergency associated with a poor prognosis, and requires prompt diagnosis and early revascularisation in order to improve outcome. The traditional management of surgical resection of necrotic bowel plus mesenteric revascularisation by surgical bypass is associated with significant morbidity and mortality. We describe the use of a combined surgical and endovascular approach, using intraoperative retrograde superior mesenteric angioplasty at the time of laparotomy. Four patients have been treated by this combined technique with three surviving, although one subsequently required an open surgical revascularisation procedure.

Epidemiology of major paediatric trauma.

In Europe injury is the leading cause of death in those aged between 1 and 14 years. In Ireland over 800,000 people are aged less than 14 years. There is currently no national trauma register to collect data on the morbidity and mortality associated with major trauma in the paediatric population in Ireland. We prospectively collected data on 153 patients admitted to our hospital with major trauma. There were 99 males and 54 females. The majority of patients were transported by ambulance (n= 138). Road traffic accidents (n=69) and thermal injuries (n=49) represented the majority of admissions. 68% (n=47) of the vehicle occupants in this study were either unrestrained or incorrectly restrained. Most patients (n=133) had an in patient stay of <50 days, with only 4 patients staying >100 days. 14 patients died. A paediatric trauma register as well as a level 1 paediatric trauma centre are required in Ireland.

Management of vascular birthmarks: review of a multidisciplinary clinic.

Vascular birthmarks comprise a diverse group of congenital lesions and represent a significant cosmetic and functional burden for patients. They remain a diagnostic and management challenge for physicians due to their extremely variable clinical presentation and often complex anatomical associations. As each type of vascular lesion has a treatment program individual to it, optimal functional and cosmetic outcomes require accurate diagnosis. Primary physicians readily diagnose and manage uncomplicated lesions, such as isolated haemangiomas and innocuous capillary malformations. However, given the complexity and relative rarity of many other vascular birthmarks, specialised multidisciplinary clinics are central to their management. In this review, we present our experience regarding the diagnostic range of vascular anomalies, associated symptomatology, and management of patients with vascular birthmarks attending the multidisciplinary Joint Vascular Birthmark Clinic at Our Lady's Children's Hospital, Crumlin. Vascular tumours represented 57% of cases reviewed, malformations accounting for 43%. Of patients not previously seen at the JVBC or by any of the individual consultants, the initial or referring diagnosis was incorrect in 42%. Significantly, 62% of vascular malformations were assigned an incorrect diagnosis, highlighting the need for a specialised clinic.

Skin secretions of Rana saharica frogs reveal antimicrobial peptides esculentins-1 and -1B and brevinins-1E and -2EC with novel insulin releasing activity.

Skin secretions of Rana saharica were evaluated for the isolation and characterisation of novel insulinotropic peptides. Crude secretions obtained from young adult frogs by mild electrical stimulation of the dorsal skin surface were purified by reverse phase HPLC yielding 80 fractions. In acute 20-min incubations with glucose responsive BRIN-BD11 cells, fractions 36-43, 46-54 and 57-63 significantly stimulated insulin release by 2- to 8-fold compared with 5.6 mM glucose alone. Pooled fractions in the latter two bands were rechromatographed to reveal 9 homogenous peaks, which elicited significant 1.3- to 3.5-fold increases in insulin release (P < 0.05). Structural analysis of the most potent non-toxic peptides was performed by mass spectrometry and automated Edman degradation. This revealed four major insulin-releasing peaks with molecular masses of 2,676.9 Da, 3,519.3 Da, 4,920.4 Da and 4,801.2 Da respectively. These peptides were found to be identical to brevinin-1E, brevinin-2EC, esculentin-1 and esculentin-1B, which belong to the group of antimicrobial peptides isolated from skin secretions of various Rana frog species. Preliminary studies on the mechanism underlying the insulinotropic actions of esculentins-1 and -1B suggested possible involvement of both cyclic AMP-protein kinase A and -C-dependent G-protein sensitive pathways. These data indicate that the skin secretions of Rana saharica frogs contain bioactive molecules with significant insulin-releasing activity. Relatives of the brevinin/esculentin peptide family merit further investigation as novel insulin secretagogues.

Are paediatric burns more common in asylum seekers? An analysis of paediatric burn admissions.

The number of asylum seekers in Ireland has increased dramatically over the last 10 years. Based on our impression that the number of children admitted to our burn unit was disproportionately represented by children of asylum seekers we performed an audit to establish (1) what proportion of admissions are from this subgroup and (2) the characteristics of their burns. All paediatric burn admissions from May 2003 to April 2004 were reviewed. Data collected from a retrospective chart review included patient demographics and details of the burn injuries. The National Census of 2002 and the Office of the Refugee Applications Commissioner were consulted for population statistics. Total burn admissions for the period were 126: Irish nationals (n=107), non-national residents (n=2), asylum seekers (n=14) and patients of unknown asylum status (n=3, excluded from study). In the asylum seeker group, the median age was 18.6 months (range 10 months-5.3 years) with the majority less than 2 years (n=11). All burns occurred in the domestic setting. Scalds accounted for 13 cases, one contact burn occurred from a hot grill. The median total body surface area burned was 5.7% (range 1.5-26%). The National Census of 2002 recorded a population of 3,917,203. With less than 12,000 asylum seekers in the country, they comprise only approximately 0.3% of the population yet they account for 11.4% of the burn patients admitted to our unit, p<0.0001. Children of asylum seekers are over-represented in our series of paediatric admissions for burns and are more likely than Irish children to sustain a burn at a younger age and in the domestic setting. This may indicate an increased risk of injury and warrants further investigation.

Clinical pharmacology of oral and i.v. N-methylformamide: a pharmacologic basis for lack of clinical antineoplastic activity.

N-Methylformamide (NMF) has been an agent of considerable interest to oncologists because of its broad spectrum of preclinical antitumor activity, tumor-differentiating abilities, and radiosensitizing and chemosensitizing properties. In this report, the pharmacokinetics of NMF are described, based on data from two phase I studies exploring both iv and oral routes of administration. Mean peak NMF plasma concentrations at recommended phase II doses were 0.46 mmol/L for NMF administered orally, 600 mg/m2 three times/week X 4 weeks every 6 weeks, and 2.78 mmol/L for NMF administered as a weekly iv bolus at 2,000 mg/m2 X 3 weeks every 4 weeks. These NMF concentrations were significantly lower than the concentrations that have been demonstrated to induce antineoplastic and relevant biologic effects in preclinical studies. Plasma disappearance curves were biphasic in the majority of patients; however, 25% of the curves were best fit by a monoexponential kinetic model. Mean alpha half-life and beta half-life values (+/- SE) were 10 +/- 2 and 732 +/- 93 min, respectively. Volumes of distribution for the theoretical central compartment (Vc) and at steady-state (Vss) were 13.8 +/- 1.1 L/m2 and 18.7 +/- 1.1 L/m2, respectively. The mean plasma clearance of NMF was 19.1 +/- 2.1 mL/min per square meter, and the relative contributions to parent compound disposition by respiratory and renal routes were insignificant. No metabolites were identified. Gastrointestinal absorption of oral NMF was rapid and nearly complete; oral bioavailability was calculated to be 0.87. Pharmacodynamic associations were observed between the magnitude of the area under the plasma disappearance curves and hepatotoxicity, the dose-limiting toxic effect of iv NMF, and the symptom complex of nausea, vomiting, and malaise, which precluded dose escalation of oral NMF.

Evaluation of a commercially available beneficial insect habitat for management of lepidoptera pests.

A field study was conducted in 2003 and 2004 at the Center for Environmental Farming Systems in Goldsboro, NC, to evaluate the effectiveness of a commercially available beneficial insect habitat in decreasing pest caterpillar populations in organically managed tomato, Lycopersicon esculentum Mill., plots. Six pairs of tomato plots were established and a commercial beneficial insect habitat seed mix (Peaceful Valley's Good Bug Blend) transplanted around the perimeter of treatment plots, whereas a brown-top millet, Brachiaria ramose (L.) Stapf., border was planted around control plots. Egg predation, egg parasitism by trichogrammatid wasps, and larval parasitism by braconid wasps was monitored throughout the growing season to determine whether habitat increased their activity. In both years of this study, the density of Helicoverpa zea (Boddie) and Manduca spp. eggs was not significantly different between treatment and control plots. Although parasitism was the most important component of egg mortality (19-49%), parasitism was not significantly different between habitat types. Identifiable predation was a minor component (3-9%) of egg fate; it is possible that unidentified predation may be part of the approximately 35-52% of eggs that met unknown fates. Larval parasitism levels ranged from approximately 10 to 90% but was not significantly influenced by the presence of beneficial insect habitat in either year of the study. These results demonstrate that natural enemy activity in organic tomatoes was not amplified, and pest populations were not reduced by the presence of a commercially available beneficial insect habitat.

Detection of individual human neutrophil alpha-defensins (human neutrophil peptides 1, 2 and 3) in unfractionated gingival crevicular fluid--a MALDI-MS approach.

The role of antimicrobial peptides is particularly important in the oral cavity where there is constant challenge by microorganisms. The alpha-defensins are a group of cationic peptides that comprise 30-50% of the total protein in azurophilic granules of human neutrophils. They include the human neutrophil peptides (HNP) 1, 2 and 3 which have almost identical amino acid sequences but differ in their biological activities. The amino acid sequence similarities of the defensins have made it difficult to unequivocally determine the presence of individual defensins using antibody-based techniques. However, by virtue of their cationic nature we postulated that the defensins would fly particularly well in mass spectrometry and that this characteristic would allow facile identification of individual HNPs in unfractionated gingival crevicular fluid (GCF) from periodontitis patients and healthy controls. Although there was variability in levels of defensins detected in periodontal health and disease, HNP-1 was always identified as the major peak in the triad and HNP-3 as the minor peak, lending support to the hypothesis that HNP-2 may arise by post-translational proteoyltic cleavage of HNP-3 rather than HNP-1. The finding that the defensins were more abundant in a higher proportion of the healthy sites studied could be linked to a more intact defensin barrier in periodontal health.

The first reported outbreak of equine herpesvirus myeloencephalopathy in New Zealand.

CASE HISTORY AND CLINICAL FINDINGS: On 9 January 2014 (Day 0) a mare from a stud farm in the Waikato region presented with urinary incontinence without pyrexia. Over the following 33 days 15 mares were clinically affected with neurological signs. All but one mare had a foal at foot. The most commonly observed clinical signs were hind limb paresis and ataxia. In some cases recumbency occurred very early in the course of disease and seven mares were subject to euthanasia for humane reasons. LABORATORY FINDINGS: Equid herpesvirus (EHV) type 1 was detected using PCR in various tissues collected post mortem from two mares with neurological signs. DNA sequencing data from the DNA polymerase gene of the virus showed a nucleotide transition at position 2254, a mutation encoding amino acid D752 that is highly associated with the neuropathogenic genotype of EHV-1. In total 12/15 mares were confirmed positive for EHV-1 on PCR. Results from a virus neutralisation test and ELISA on paired serum samples, and PCR on whole blood and nasal swabs, indicated that of four paddocks in a high-risk area where a cluster of cases had occurred, 20/21 (95%) horses were likely to have been exposed or were confirmed infected with EHV-1. Subsequent to the outbreak two mares aborted, one at 9 months and one at 10 months of gestation. The cause of abortion was confirmed as EHV-1 with the same genotype as that involved in the outbreak. DIAGNOSIS: Equine herpesvirus myeloencephalopathy. CLINICAL RELEVANCE: The outbreak described shows the considerable impact that can occur in outbreaks of equine herpesvirus myeloencephalopathy in New Zealand. Early biosecurity controls not only reduced the effect on the farm but mitigated the potential for the virus to spread to other horse enterprises.

Continuous interleukin-2 and lymphokine-activated killer cells for advanced cancer: a National Biotherapy Study Group trial.

We conducted a multicenter, phase II trial of continuous-infusion recombinant interleukin-2 (rIL-2) and lymphokine-activated killer (LAK) cells. Patients had advanced cancer, measurable disease, and a good performance level. Treatment included a 5-day continuous infusion of 18 x 10(6) IU/m2/d of rIL-2 followed by 1 day of rest, 4 days of leukapheresis to collect cells for in vitro augmentation of cellular cytotoxicity, and 5 more days of rIL-2 infusion with reinfusion of LAK cells for 3 successive days. Therapy was repeated after 2 weeks. There were 117 patients enrolled: 63% were males, with a median age of 51 years. Eighty-two percent were managed in oncology units, and 18% were in intensive care units. Six patients died within 1 month of initiating therapy. In renal cell carcinoma, the response rate was one of 31 patients (3%), with a median survival of 10.7 months. In melanoma, the response rate was four of 33 patients (12%), with a median survival of 6.1 months. For all other histologies, response rate was three of 53 patients (5%), with a median survival of 7.4 months. All responders were asymptomatic when therapy was initiated. This trial confirms the feasibility of administering continuous rIL-2 and LAK cells outside the intensive care unit environment. Antitumor activity in melanoma was similar to that seen in multicenter trials of bolus rIL-2 and LAK cells. Activity in renal cell cancer was disappointing.

Randomized, double-blind, placebo-controlled trial to evaluate the hematopoietic growth factor PIXY321 after moderate-dose fluorouracil, doxorubicin, and cyclophosphamide in stage II and III breast cancer.

PURPOSE: To measure the effect of PIXY321 (granulocyte-macrophage colony-stimulating factor/interleukin-3 S. cerevisiae fusion protein) on the incidence, duration, and complications of neutropenia and thrombocytopenia after moderate-dose fluorouracil 600 mg/m(2), doxorubicin 60 mg/m(2), and cyclophosphamide 750 mg/m(2) (FAC) chemotherapy in patients with stage II and III breast cancer. PATIENTS AND METHODS: In this multicenter, randomized, double-blind placebo-controlled trial, 71 women were to receive four 21-day cycles of treatment with moderate-dose FAC chemotherapy by short intravenous infusion on day 1, followed by either placebo or PIXY321 (375 microg/m(2) subcutaneously twice a day) on days 3 to 15. All patients were to receive prophylactic oral ciprofloxacin when the absolute neutrophil count was less than 1,000/microL. RESULTS: PIXY321 significantly reduced the incidence and duration of grade 3 and grade 4 neutropenia in cycles 1 and 2 and the duration of grade 3 neutropenia in cycles 1 through 4. In cycles 3 and 4, grade 3 thrombocytopenia was significantly more common with PIXY321 (P <.05). Two patients, both in the PIXY321 group, required platelet transfusions. Fever and hospitalization for intravenous antibiotics were significantly more common in the PIXY321 group during cycle 1 only. More patients in the PIXY321 group achieved hematologic recovery by day 22 in cycles 1 through 3, and time to recovery was significantly shorter with PIXY321 in all cycles. FAC dose intensity was roughly 2% higher in the PIXY321 group (P = NS). Nonhematologic events of any intensity occurring with significantly greater overall frequency in the PIXY321 group included injection-site reactions, fever, chills, abdominal pain, and arthralgia. No patient died on study or within 30 days of her last dose of study drug. CONCLUSION: PIXY321 decreased the incidence and duration of FAC-induced grade 3 and 4 neutropenia in cycles 1 and 2 and significantly shortened the time to hematologic recovery in all cycles. However, it produced more systemic toxicity as well as thrombocytopenia in cycles 3 and 4.

Phenotypic and genetic analysis of Clock, a new circadian rhythm mutant in Drosophila melanogaster.

Clock is a semidominant X-linked mutation that results in shortening the period of Drosophila melanogaster's free-running locomotor activity rhythm from ca. 24.0 to ca. 22.5 hr. This mutation similarly shortened the phase response curve, determined by resetting activity rhythms with light pulses. Eclosion peaks for Clk cultures were separated by only 22.5 hr instead of the normal 24 hr. Clk was mapped close to, but separable from, another rhythm mutation--period01--by recombination. The estimated distance between these two mutations was short enough to suggest that Clk could be a per allele. If this is the case, the new mutant is unique in that it, unlike other per variants, is associated with essentially normal 1-min courtship song rhythms when Clk is expressed in males. Also, the new rhythm variant could not, in contrast to a short-period per mutation, have its effects on free-running activity rhythms uncovered by deletions. This result, and the lack of coverage of Clk's effects by duplications, suggest that it is not a simple hypomorphic or amorphic mutation.