Temperature Restriction in Entomopathogenic Bacteria.

Temperature plays an important role in bacteria-host interactions and can be a determining factor for host switching. In this study we sought to investigate the reasons behind growth temperature restriction in the entomopathogenic enterobacterium Photorhabdus. Photorhabdus has a complex dual symbiotic and pathogenic life cycle. The genus consists of 19 species but only one subgroup, previously all classed together as Photorhabdus asymbiotica, have been shown to cause human disease. These clinical isolates necessarily need to be able to grow at 37°C, whilst the remaining species are largely restricted to growth temperatures below 34°C and are therefore unable to infect mammalian hosts. Here, we have isolated spontaneous mutant lines of Photorhabdus laumondii DJC that were able to grow up to 36-37°C. Following whole genome sequencing of 29 of these mutants we identified a single gene, encoding a protein with a RecG-like helicase domain that for the majority of isolates contained single nucleotide polymorphisms. Importantly, provision of the wild-type allele of this gene in trans restored the temperature restriction, confirming the mutations are recessive, and the dominant effect of the protein product of this gene. The gene appears to be part of a short three cistron operon, which we have termed the Temperature Restricting Locus (TRL). Transcription reporter strains revealed that this operon is induced upon the switch from 30 to 36°C, leading to replication arrest of the bacteria. TRL is absent from all of the human pathogenic species so far examined, although its presence is not uniform in different strains of the Photorhabdus luminescens subgroup. In a wider context, the presence of this gene is not limited to Photorhabdus, being found in phylogenetically diverse proteobacteria. We therefore suggest that this system may play a more fundamental role in temperature restriction in diverse species, relating to as yet cryptic aspects of their ecological niches and life cycle requirements.

Polyproline as a Minimal Antifreeze Protein Mimic That Enhances the Cryopreservation of Cell Monolayers.

Tissue engineering, gene therapy, drug screening, and emerging regenerative medicine therapies are fundamentally reliant on high-quality adherent cell culture, but current methods to cryopreserve cells in this format can give low cell yields and require large volumes of solvent "antifreezes". Herein, we report polyproline as a minimum (bio)synthetic mimic of antifreeze proteins that is accessible by solution, solid-phase, and recombinant methods. We demonstrate that polyproline has ice recrystallisation inhibition activity linked to its amphipathic helix and that it enhances the DMSO cryopreservation of adherent cell lines. Polyproline may be a versatile additive in the emerging field of macromolecular cryoprotectants.

Propagation and some physiological effects of Citrus bark cracking viroid and Apple fruit crinkle viroid in multiple infected hop (Humulus lupulus L.).

The hop metabolome important for the brewing industry and for medical purposes is endangered worldwide due to multiple viroid infections affecting hop physiology. Combinatorial biolistic hop inoculation with Citrus bark cracking viroid (CBCVd), Apple fruit crinkle viroid (AFCVd), Hop latent viroid, and Hop stunt viroid (HSVd) showed a low CBCVd compatibility with HSVd, while all other viroid combinations were highly compatible. Unlike to other viroids, single CBCVd propagation showed a significant excess of (-) over (+) strands in hop, tomato, and Nicotiana benthamiana, but not in citruses. Inoculation of hop with all viroids led to multiple infections with unstable viroid levels in individual plants in the pre- and post-dormancy periods, and to high plant mortality and morphological disorders. Hop isolates of CBCVd and AFCVd were highly stable, only minor quasispecies were detected. CBCVd caused a strong suppression of some crucial mRNAs related to the hop prenylflavonoid biosynthesis pathway, while AFCVd-caused effects were moderate. According to mRNA degradome analysis, this suppression was not caused by a direct viroid-specific small RNA-mediated degradation. CBCVd infection led to a strong induction of two hop transcription factors from WRKY family and to a disbalance of WRKY/WDR1 complexes important for activation of lupulin genes.

Viroid quasispecies revealed by deep sequencing.

Viroids are non-coding single-stranded circular RNA molecules that replicate autonomously in infected host plants causing mild to lethal symptoms. Their genomes contain about 250-400 nucleotides, depending on viroid species. Members of the family Pospiviroidae, like the Potato spindle tuber viroid (PSTVd), replicate via an asymmetric rolling-circle mechanism using the host DNA-dependent RNA-Polymerase II in the nucleus, while members of Avsunviroidae are replicated in a symmetric rolling-circle mechanism probably by the nuclear-encoded polymerase in chloroplasts. Viroids induce the production of viroid-specific small RNAs (vsRNA) that can direct (post-)transcriptional gene silencing against host transcripts or genomic sequences. Here, we used deep-sequencing to analyze vsRNAs from plants infected with different PSTVd variants to elucidate the PSTVd quasipecies evolved during infection. We recovered several novel as well as previously known PSTVd variants that were obviously competent in replication and identified common strand-specific mutations. The calculated mean error rate per nucleotide position was less than [Formula: see text], quite comparable to the value of [Formula: see text] reported for a member of Avsunviroidae. The resulting error threshold allows the synthesis of longer-than-unit-length replication intermediates as required by the asymmetric rolling-circle mechanism of members of Pospiviroidae.

Ovarian ectopic pregnancy: aetiology, diagnosis, and challenges in surgical management.

Primary ovarian ectopic pregnancy (OEP) remains a rare but important type of ectopic pregnancy which is normally diagnosed at surgery. The aetiology is unknown. We have found fertility treatments (18.1%) and intrauterine contraceptive devices (19.3%) remain important associated risk factors. We undertook a world literature review of all cases since 1995. We found that 52% of cases were managed laparoscopically, which is less than the reported UK rate of all ectopics, of 85.9%. Over that time period, eight patients were treated medically with a success rate of 50%. While the original 1878 Spiegelberg's criterion still holds for the diagnosis, the addition of modern ultrasound scanning has made a radiological diagnosis possible. In a few cases, the management of OEP remains similar to that of a tubal ectopic pregnancy with the same challenges. Laparoscopy with ovarian sparing in the stable woman is today the surgical treatment of choice.

The neurocognitive connection between physical activity and eating behaviour.

As obesity rates increase worldwide, healthcare providers require methods to instill the lifestyle behaviours necessary for sustainable weight loss. Designing effective weight-loss interventions requires an understanding of how these behaviours are elicited, how they relate to each other and whether they are supported by common neurocognitive mechanisms. This may provide valuable insights to optimize existing interventions and develop novel approaches to weight control. Researchers have begun to investigate the neurocognitive underpinnings of eating behaviour and the impact of physical activity on cognition and the brain. This review attempts to bring these somewhat disparate, yet interrelated lines of literature together in order to examine a hypothesis that eating behaviour and physical activity share a common neurocognitive link. The link pertains to executive functions, which rely on brain circuits located in the prefrontal cortex. These advanced cognitive processes are of limited capacity and undergo relentless strain in the current obesogenic environment. The increased demand on these neurocognitive resources as well as their overuse and/or impairment may facilitate impulses to over-eat, contributing to weight gain and obesity. This impulsive eating drive may be counteracted by physical activity due to its enhancement of neurocognitive resources for executive functions and goal-oriented behaviour. By enhancing the resources that facilitate 'top-down' inhibitory control, increased physical activity may help compensate and suppress the hedonic drive to over-eat. Understanding how physical activity and eating behaviours interact on a neurocognitive level may help to maintain a healthy lifestyle in an obesogenic environment.

Autologous blood collection and transfusion in cats undergoing partial craniectomy.

OBJECTIVE: To describe the procedure for autologous blood donation and associated complications in cats undergoing partial craniectomy for mass removal. DESIGN: Prospective case series. ANIMALS: 15 cats with intracranial mass confirmed by computed tomographic scan, no evidence of renal failure, and PCV > or = 22%. PROCEDURE: One unit (60 ml) of blood was collected and stored 7 to 17 days before surgery and transfused during the perioperative period if needed. The PCV was measured before donation, before surgery, during surgery, and after surgery to assess effect of donation on PCV before surgery and effect of transfusion on PCV after surgery. Cats were evaluated for donation complications, iatrogenic anemia, and adverse reactions associated with administration of autologous blood. RESULTS: Complications associated with phlebotomy were not detected. Fifteen cats underwent partial craniectomy 7 to 17 days after blood donation; all had histologic confirmation of meningioma by examination of tissue obtained at surgery. Eleven cats received autologous blood transfusions. None of the cats received allogeneic blood transfusions. Transfusion reactions were not observed. Subclinical iatrogenic anemia was detected in 3 cats. Two cats were considered to have received excessive transfusion, and 3 cats received inadequate transfusion. All cats undergoing partial craniectomy were discharged from the hospital and were alive > 6 months after surgery. CONCLUSIONS AND CLINICAL RELEVANCE: Autologous blood donation before surgery was considered safe for cats undergoing partial craniectomy for resection of meningioma. The only complication observed was iatrogenic anemia. The procedure contributed to blood conservation in our hospital.

Epidemiology of HIV and AIDS. A retrospective look.

This retrospective look at literature and information regarding the epidemiology of HIV and its progression to AIDS is based on many worldwide sources to determine with some certainty the true severity of the epidemic. The purpose of this article is to permit the reader to become more informed concerning the epidemic, based on a global outlook.

PIP: Global survey estimates of HIV/AIDS suggest an overwhelming increase in the severity of the epidemic. According to UN survey estimates, over 3.1 million people were infected with HIV in 1996, with 8500 new cases occurring per day. About 5 million adults and 1.5 million children have died since the beginning of the epidemic, and this proportion is continually increasing. Substantial increases in the HIV prevalence rate are observed in Vietnam, Thailand, Europe, the US, Australia and sub-Saharan Africa. The majority of these infections is transmitted through unprotected sexual intercourse as well as intravenous drug use. While people of reproductive age are the most common victims of HIV/AIDS, children are not spared from contracting the disease. Studies have shown that this deadly disease can be controlled and prevented through early identification and initiation of antiretroviral therapy, use of prophylactic medications to prevent secondary AIDS-related diseases, as well as vigorous implementation of prevention activities such as community outreach and needle-exchange programs. Since the clinical status of most persons infected with HIV has not yet progressed to AIDS, AIDS prevalence underestimates the total number of people with HIV in need of related services and further highlights the need for early detection and prevention methods.

Results of craniotomy for the treatment of cerebral meningioma in 42 cats.

Forty-two cats underwent craniotomy for removal of a meningioma between 1985 and 1991. Median duration of clinical signs before examination was 1.25 months. All cats had inappropriate demeanor: 48% were dull and 38% were lethargic. Neurological deficits included impaired vision in 93%, paresis in 83%, and seizures in 19%. Computed tomography (CT) showed solitary masses in 86% and multiple masses in 14%. Intraoperative complications included hemorrhage and difficulty excising deep or adherent masses. Anemia in 13 of 42 cats was the most common immediate postoperative complication. Ten of 42 cats had no improvement or a more severe neurological status after surgery. Eight of 42 cats died immediately after surgery; 6 of these were anemic. Of the cats that survived the immediate postoperative period, evaluation 10 to 14 days after surgery showed that 97% (33 of 34) were alert and 79% (27 of 34) had returned to normal behavior. Neurological deficits, except for vision impairment, had resolved in most cats. The duration of follow-up varied from 1.3 months to 55.1 months. Ten cats developed neurological abnormalities from 1 month to 44.2 months after surgery; of these, 6 had tumor recurrence or new growth confirmed by CT scan or necropsy. Overall survival was 71% at 6 months, 66% at 1 year, and 50% at 2 years. Age of cat and location of tumor did not significantly affect survival (P = .1034 and .1851, respectively). There were too few precise measurements of tumor size to make a valid statistical comparison of the effect of size on survival. Location or presence of multiple tumors did not affect final outcome.(ABSTRACT TRUNCATED AT 250 WORDS)

Idiopathic hypoparathyroidism in five cats.

Idiopathic hypoparathyroidism was diagnosed in five young to middle-aged cats of mixed breeding. Three of the cats were male and two were female. Historic signs included lethargy (n = 5), anorexia (n = 5), muscle tremors (n = 4), weakness (n = 4), generalized seizures (n = 3), ataxia (n = 3), mental dullness or disorientation (n = 3), panting (n = 2), pruritus (n = 1), ptyalism (n = 1) and dysphagia (n = 1). Weakness (n = 4), dehydration (n = 2), cataracts (n = 2), hypothermia (n = 1), and bradycardia (n = 1) were found on physical examination. Results of electrocardiography revealed a prolonged Q-T interval in two cats. Results of initial laboratory tests revealed profound hypocalcemia and severe hyperphosphatemia with normal renal function. The diagnosis of hypoparathyroidism was made on the basis of the history, clinical signs, and results serum biochemical testing (i.e., severe hypocalcemia and hyperphosphatemia); in two cats, the diagnosis was also confirmed by histologic examination of parathyroid glands. Initial treatment included intravenous administration of 10% calcium gluconate and oral administration of large loading doses of calcium and vitamin D (dihydrotachysterol). Successful long-term management with dihydrotachysterol and calcium was achieved in all cats. The final dosage of dihydrotachysterol required to maintain normocalcemia in the five cats ranged from 0.004 to 0.04 mg/kg/day (mean = 0.015 mg/kg/day). Long-term calcium supplementation was given to three of the cats in dosages ranging from 29 to 53 mg/kg/day (mean = 42 mg/kg/day) of elemental calcium. One cat died after 28 months of therapy from widely metastatic hemangiosarcoma; the other three cats are still alive and well after 5 to 37 months of treatment.

The differential diagnosis of disc disease.

The differentiation of disc disease from other diseases that result in signs related to varying degrees of proprioceptive ataxia, paresis to paralysis, digital hypesthesia to anesthesia, and spinal hyperpathia is a common problem facing the veterinarian. Classification of the differential diagnoses of disc disease based on neuroanatomic localization facilitates diagnosis because each category shares historical, clinical, and neurologic examination features. These features, as well as the distinguishing ones used in the differential diagnosis of disc disease, will be reviewed.

Myasthenia gravis in the cat.

Myasthenia gravis (MG) was diagnosed in four cats--one had an apparently congenital form and three had the acquired autoimmune form. All four cats were examined because of episodes of weakness including gait abnormalities, voice change, neck ventroflexion, and regurgitation. Palpebral reflexes were absent in all cats. Administration of edrophonium chloride resulted in transient resolution of clinical signs in all four cats. Three cats were tested for the presence of serum autoantibodies against acetylcholine receptor (AChR) by radioimmunoassay. Two cats with acquired MG had anti-AChR antibody titers of 10.5 and 96.8 nmol/l (normal, less than or equal to 0.03 nmol/l). Antibodies were not detected in the cat with presumptive congenital MG. All four cats were treated with pyridostigmine bromide. Two cats with acquired MG were euthanatized because of clinical deterioration. The third cat with acquired MG has been asymptomatic since 2 months after diagnosis. The cat with presumed congenital MG is alive 3 years after diagnosis.

Evaluation of two reagent strips and three reflectance meters for rapid determination of blood glucose concentrations.

We evaluated three reflectance meters (Accu-Chek II, Glucometer II, and Glucoscan 2000) and two reagent strips (Chemstrip bG and Glucostix) for accuracy and precision in determining blood glucose concentrations in the dog. To evaluate accuracy, we compared results of blood glucose determinations performed on 95 samples using the various strips and meters vs. the glucose concentrations obtained using the glucose-oxidase method on a Beckman Glucose Analyzer. Accuracy was evaluated statistically using least squares regression analysis. To evaluate precision, samples in various ranges of blood glucose concentration were tested repeatedly (20 times within a 1-hour period) on the same reflectance meter. Coefficient of variation (CV) was determined to evaluate reproducibility of results. Overall, there were significant correlations (P less than 0.001) between the laboratory glucose values and the blood glucose concentrations obtained with Chemstrip bG (r = 0.976), Glucostix (r = 0.904), Accu-Chek II (r = 0.986), Glucometer II (r = 0.911) and Glucoscan 2000 (r = 0.944). In the precision study, all three meters had excellent CVs in the normal range (3.6% to 4.9%). However, Accu-Chek II was found to be more precise in the hypoglycemic and hyperglycemic ranges (3.6% and 2.6%, respectively) than either Glucometer II (8.8% and 5.4%) or Glucoscan 2000 (7.8% and 8.2%). The results of this study indicate that all of the meters and reagent strips tested are highly accurate in determining blood glucose concentrations in the dog. However, both in terms of accuracy and reproducibility of results, Accu-Chek II and Chemstrip bG, gave the highest correlation coefficients and, as such, are probably of the greatest clinical value.