Reactivation of transcription from a vaccinia virus early promoter late in infection.

We have studied the kinetics of RNA synthesis from the vaccinia virus 7,500-molecular-weight gene (7.5K gene) which is regulated by early and late promoters arranged in tandem. Unexpectedly, after a first burst of RNA synthesis early in infection, transcription was reactivated late in infection. Reactivation was not dependent on the location of the promoter in the genome or on the presence of the upstream late regulatory sequences. The mRNA synthesized from the reactivated promoter in the late phase had the same 5' and 3' ends as the molecules transcribed in the early phase. Interestingly, these molecules were efficiently translated despite the absence of the poly(A) leader characteristic of late mRNAs. Reactivation appears to be dependent on virus assembly since it is prevented by rifampin, a specific inhibitor of morphogenesis. Finally, analysis of various other early genes showed that reactivation is not unique to the 7.5K early promoter.

Circadian expression of H,K-ATPase type 2 contributes to the stability of plasma K⁺ levels.

Maintenance by the kidney of stable plasma K(+) values is crucial, as plasma K(+) controls muscle and nerve activity. Since renal K(+) excretion is regulated by the circadian clock, we aimed to identify the ion transporters involved in this process. In control mice, the renal mRNA expression of H,K-ATPase type 2 (HKA2) is 25% higher during rest compared to the activity period. Conversely, under dietary K(+) restriction, HKA2 expression is ∼40% higher during the activity period. This reversal suggests that HKA2 contributes to the circadian regulation of K(+) homeostasis. Compared to their wild-type (WT) littermates, HKA2-null mice fed a normal diet have 2-fold higher K(+) renal excretion during rest. Under K(+) restriction, their urinary K(+) loss is 40% higher during the activity period. This inability to excrete K(+) "on time" is reflected in plasma K(+) values, which vary by 12% between activity and rest periods in HKA2-null mice but remain stable in WT mice. Analysis of the circadian expression of HKA2 regulators suggests that Nrf2, but not progesterone, contributes to its rhythmicity. Therefore, HKA2 acts to maintain the circadian rhythm of urinary K(+) excretion and preserve stable plasma K(+) values throughout the day.

New clinical practice guideline on enteral feeding in very low birth wieght infants; second part

INTRODUCTION: The nutrition of very low birth weight (VLBW) infants is aimed at promoting a similar growth to that occurring in the uterus. However, in practice this is difficult to achieve and extrauterine growth restriction is frequent. The current tendency is to avoid this restriction by means of early parenteral and enteral nutrition. Nonetheless, uncertainty about many of the practices related with nutrition has resulted in a great variation in the way it is undertaken. In 2009 and 2011 in our hospital there was an unexpected increase in necrotizing enterocolitis. To check to see wether our nutrition policy was involved, we underlook a systematic review and drewup clinical practice guidelines (CPG) about enteral feeding in VLBW infants. New considerations about the duration of the fortification and the use of probiotics have led to an update of these CPG. METHODS: A total of 21 clinical questions were designed dealing with the type of milk, starting age, mode of administration, rate and volume of the increments, fortification, use of probiotics and protocol. Afete conducting a systematic search of the available evidence, the information was contrasted and summarized in order to draw up the recommendations. The quality of the evidence and the strength of the recommendations were determined from the SIGN scale. COMMENT: These CPG aim to help physicians in their decision making. The protocolized application of wellproven measurements reduces the variation in clinical practice and improves results.

Hypersensitivity pneumonitis secondary to lovebirds: a new cause of bird fancier's disease.

Hypersensitivity pneumonitis (HP) is an immunologically mediated lung disease due to the repetitive inhalation of antigens. Most new cases arise from residential exposures, notably to birds, and are thus more difficult to recognise. The present authors report a 59-yr-old male who complained of dyspnoea and cough while being treated with amiodarone. Pulmonary function tests revealed restriction and obstruction with low diffusing lung capacity for carbon monoxide and partial pressure of oxygen. A high-resolution computed tomography chest scan and bronchoalveolar lavage showed diffuse bilateral ground-glass attenuation and lymphocytic alveolitis, respectively. Initial diagnosis was amiodarone pulmonary toxicity, but because of a rapidly favourable evolution, this diagnosis was questioned. A careful environmental history revealed a close contact with lovebirds shortly before the onset of symptoms. Precipitins were strongly positive against lovebird droppings, but were negative against other avian antigens. The patient was diagnosed with hypersensitivity pneumonitis to lovebirds. Avoidance of lovebirds and steroid treatment led to rapid improvement. The present observation identifies a new causative agent for hypersensitivity pneumonitis and highlights the importance of a thorough environmental history and of searching for precipitins against antigens directly extracted from the patient's environment. These two procedures should allow a more precise classification of some cases of pneumonitis, and thus might avoid progression of active undiagnosed hypersensitivity pneumonitis to irreversible fibrosis or emphysema.

Molecular characterisation and disease severity of leptospirosis in Sri Lanka

Leptospirosis is a re-emerging zoonotic disease all over the world, important in tropical and subtropical areas. A majority of leptospirosis infected patients present as subclinical or mild disease while 5-10% may develop severe infection requiring hospitalisation and critical care. It is possible that several factors, such as the infecting serovar, level of leptospiraemia, host genetic factors and host immune response, may be important in predisposition towards severe disease. Different Leptospirastrains circulate in different geographical regions contributing to variable disease severity. Therefore, it is important to investigate the circulating strains at geographical locations during each outbreak for epidemiological studies and to support the clinical management of the patients. In this study immunochromatography, microscopic agglutination test and polymerase chain reaction were used to diagnose leptospirosis. Further restriction fragment length polymorphism and DNA sequencing methods were used to identify the circulating strains in two selected geographical regions of Sri Lanka.Leptospira interrogans, Leptospira borgpeterseniiandLeptospira kirschneristrains were identified to be circulating in western and southern provinces. L. interroganswas the predominant species circulating in western and southern provinces in 2013 and its presence was mainly associated with renal failure.

The effect of host plant and isolation on the genetic structure of phytophagous insects: A preliminary study on a bruchid beetle

Genetic differentiation is a consequence of the combination of drift and restriction in gene flow between populations due to barriers to dispersal, or selection against individuals resulting from inter-population matings In phytophagous insects, local adaptation to different kinds of host plants can sometimes lead to reproductive isolation and thus to genetic structuring, or even to speciation Acanthoscelides. obtectus Say is a bean bruchid specialized on beans of the Phaseolus vulgaris group, attacking both wild and domesticated forms of P vulgaris., and P coccineus This study reveals that the genetic structure of populations of this bruchid is explained mainly by their geographical location and is not related to a particular kind (wild or domesticated) of bean In contrast, the species of bean might have led, to some extent, to genetic structuring in these bruchids, although our sampling is too limited to address such process unambiguously. If confirmed, it would corroborate preliminary results found for the parasitoid species that attack Acanthoscelides species, which might show a genetic structure depending on the species of host plant

Polymerase chain reaction detection of LeishmaniaDNA in skin biopsy samples in Sri Lanka where the causative agent of cutaneous leishmaniasis is Leishmania donovani

Leishmania donovani is the known causative agent of both cutaneous (CL) and visceral leishmaniasis in Sri Lanka. CL is considered to be under-reported partly due to relatively poor sensitivity and specificity of microscopic diagnosis. We compared robustness of three previously described polymerase chain reaction (PCR) based methods to detectLeishmania DNA in 38 punch biopsy samples from patients presented with suspected lesions in 2010. Both, Leishmaniagenus-specific JW11/JW12 KDNA and LITSR/L5.8S internal transcribed spacer (ITS)1 PCR assays detected 92% (35/38) of the samples whereas a KDNA assay specific forL. donovani (LdF/LdR) detected only 71% (27/38) of samples. All positive samples showed a L. donovani banding pattern upon HaeIII ITS1 PCR-restriction fragment length polymorphism analysis. PCR assay specificity was evaluated in samples containing Mycobacterium tuberculosis, Mycobacterium leprae, and human DNA, and there was no cross-amplification in JW11/JW12 and LITSR/L5.8S PCR assays. The LdF/LdR PCR assay did not amplify M. leprae or human DNA although 500 bp and 700 bp bands were observed in M. tuberculosis samples. In conclusion, it was successfully shown in this study that it is possible to diagnose Sri Lankan CL with high accuracy, to genus and species identification, using Leishmania DNA PCR assays.

Peritalar dislocations: a retrospective study of 18 cases.

The purpose of this study is to retrospectively evaluate 18 consecutive cases of peritalar dislocations referred to our department during a period of 25 years and to delineate the factors influencing long-term prognosis. There were 13 (73%) medial and 5 (27%) lateral dislocations. Six patients (33%) suffered an open injury, including 2 of 13 (15%) medial and 4 of 5 (80%) lateral dislocations. Associated fractures involving the hindfoot or forefoot were noted in 7 feet, including 3 of 5 lateral dislocation cases. Reduction was accomplished under general anesthesia; in no case was open reduction necessary. In 4 of 6 open injuries with associated fractures, temporary fixation with Kirschner wires was performed. Patients were immobilized in a plaster cast for 4 weeks, or for 6 weeks in the presence of fracture, followed by weightbearing as tolerated. At a mean follow-up of 10.2 years (range, 4 to 26 years), 10 patients (56%) showed excellent results; all had sustained a closed medial low-energy dislocation. There were 3 cases (17%) with fair results and 5 cases (28%) with poor results. Forty-five percent of patients showed a restriction of activity, a reduction of subtalar range of motion, and moderate or severe radiographic signs of hindfoot degenerative arthritis. There were no cases of talar avascular necrosis, and in no case was secondary surgery necessary. Lateral dislocation and open medial dislocations with concomitant fractures showed a greater potential for poor prognosis. The results were independent from period of cast immobilization, suggesting that 4 to 6 weeks of immobilization provides acceptable long-term results.

Genetic diversity of Cryptosporidium identified in clinical samples from cities in Brazil and Argentina

The identification and characterisation of Cryptosporidiumgenotypes and subtypes are fundamental to the study of cryptosporidiosis epidemiology, aiding in prevention and control strategies. The objective was to determine the genetic diversity ofCryptosporidium in samples obtained from hospitals of Rio de Janeiro, Brazil, and Buenos Aires, Argentina. Samples were analysed by microscopy and TaqMan polymerase chain reaction (PCR) assays forCryptosporidium detection, genotyped by nested-PCR-restriction fragment length polymorphism (RFLP) analysis of the 18S rRNA gene and subtyped by DNA sequencing of the gp60 gene. Among the 89 samples from Rio de Janeiro, Cryptosporidium spp were detected in 26 by microscopy/TaqMan PCR. In samples from Buenos Aires,Cryptosporidium was diagnosed in 15 patients of the 132 studied. The TaqMan PCR and the nested-PCR-RFLP detected Cryptosporidium parvum, Cryptosporidium hominis, and co-infections of both species. In Brazilian samples, the subtypes IbA10G2 and IIcA5G3 were observed. The subtypes found in Argentinean samples were IbA10G2, IaA10G1R4, IaA11G1R4, and IeA11G3T3, and mixed subtypes of Ia and IIa families were detected in the co-infections. C. hominis was the species more frequently detected, and subtype family Ib was reported in both countries. Subtype diversity was higher in Buenos Aires than in Rio de Janeiro and two new subtypes were described for the first time.

Polyphasic characterisation of Burkholderia cepaciacomplex species isolated from children with cystic fibrosis

Cystic fibrosis (CF) patients with Burkholderia cepacia complex (Bcc) pulmonary infections have high morbidity and mortality. The aim of this study was to compare different methods for identification of Bcc species isolated from paediatric CF patients. Oropharyngeal swabs from children with CF were used to obtain isolates of Bcc samples to evaluate six different tests for strain identification. Conventional (CPT) and automatised (APT) phenotypic tests, polymerase chain reaction (PCR)-recA, restriction fragment length polymorphism-recA, recAsequencing, and matrix-assisted laser desorption ionization-time of flight (MALDI-TOF) were applied. Bacterial isolates were also tested for antimicrobial susceptibility. PCR-recA analysis showed that 36 out of the 54 isolates were Bcc. Kappa index data indicated almost perfect agreement between CPT and APT, CPT and PCR-recA, and APT and PCR-recA to identify Bcc, and MALDI-TOF and recAsequencing to identify Bcc species. The recAsequencing data and the MALDI-TOF data agreed in 97.2% of the isolates. Based on recA sequencing, the most common species identified were Burkholderia cenocepacia IIIA (33.4%),Burkholderia vietnamiensis (30.6%), B. cenocepaciaIIIB (27.8%), Burkholderia multivorans (5.5%), and B. cepacia (2.7%). MALDI-TOF proved to be a useful tool for identification of Bcc species obtained from CF patients, although it was not able to identify B. cenocepacia subtypes.

Bargaining failures and merger policy

In this paper we study the optimal ex-ante merger policy in a model where merger proposals are the result of strategic bargaining among alternative candidates. We allow for firm asymmetries and, in particular, we emphasize the fact that potential synergies generated by a merger may vary substantially depending on the identity of the participating firms. The model demonstrates that, under some circumstances, relatively inefficient mergers may take place. That is, a particular merger may materialize despite the existence of an alternative merger capable of generating higher social surplus and even higher profits. Such bargaining failures have important implications for the ex-ante optimal merger policy. We show that a more stringent policy than the ex-post optimal reduces the scope of these bargaining failures and raises expected social surplus. We use a bargaining model that is flexible, in the sense that its strategic structure does not place any exogenous restriction on the endogenous likelihood of feasible mergers.

CCR2 and CCR5 genes polymorphisms in women with cervical lesions from Pernambuco, Northeast Region of Brazil: a case-control study

Polymorphisms in chemokine receptors play an important role in the progression of cervical intraepithelial neoplasia (CIN) to cervical cancer (CC). Our study examined the association of CCR2-64I (rs1799864) andCCR5-Δ32 (rs333) polymorphisms with susceptibility to develop cervical lesion (CIN and CC) in a Brazilian population. The genotyping of 139 women with cervical lesions and 151 women without cervical lesions for the CCR2-64I and CCR5-Δ32 polymorphisms were performed using polymerase chain reaction-restriction fragment length polymorphism. The individuals carrying heterozygous or homozygous genotypes (GA+AA) for CCR2-64I polymorphisms seem to be at lower risk for cervical lesion [odds ratio (OR) = 0.37, p = 0.0008)]. The same was observed for the A allele (OR = 0.39, p = 0.0002), while no association was detected (p > 0.05) with CCR5-Δ32 polymorphism. Regarding the human papillomavirus (HPV) type, patients carrying the CCR2-64Ipolymorphism were protected against infection by HPV type 16 (OR = 0.35, p = 0.0184). In summary, our study showed a protective effect ofCCR2-64I rs1799864 polymorphism against the development of cervical lesions (CIN and CC) and in the susceptibility of HPV 16 infection.

New clinical practice guideline on enteral feeding in very low birth weight infants; first part.

The nutrition of very low birth weight (VLBW) infants is aimed at promoting a similar growth to that occurring in the uterus. However, in practice this is difficult to achieve and extrauterine growth restriction is frequent. The current tendency is to avoid this restriction by means of early parenteral and enteral nutrition. Nonetheless, uncertainty about many of the practices related with nutrition has resulted in a great variation in the way it is undertaken. In 2009 and 2011 in our hospital there was an unexpected increase in necrotizing enterocolitis. To check to see whether our nutrition policy was involved, we undertook a systematic review and drew up clinical practice guidelines (CPG) about enteral feeding in VLBW infants. New considerations about the duration of the fortification and the use of probiotics have led to an update of these CPG. METHODS: A total of 21 clinical questions were designed dealing with the type of milk, starting age, mode of administration, rate and volume of the increments, fortification, use of probiotics and protocol. After conducting a systematic search of the available evidence, the information was contrasted and summarized in order to draw up the recommendations. The quality of the evidence and the strength of the recommendations were determined from the SIGN scale. COMMENT: These CPG aim to help physicians in their decision making. The protocolized application of well-proven measurements reduces the variation in clinical practice and improves results.

A fast-track program reduces complications and length of hospital stay after open colonic surgery.

BACKGROUND & AIMS: A fast-track program is a multimodal approach for patients undergoing colonic surgery that combines stringent regimens of perioperative care (fluid restriction, optimized analgesia, forced mobilization, and early oral feeding) to reduce perioperative morbidity, hospital stay, and cost. We investigated the impact of a fast-track protocol on postoperative morbidity in patients after open colonic surgery. METHODS: A randomized trial of patients in 4 teaching hospitals in Switzerland included 156 patients undergoing elective open colonic surgery who were assigned to either a fast-track program or standard care. The primary end point was the 30-day complication rate. Secondary end points were severity of complications, hospital stay, and compliance with the fast-track protocol. RESULTS: The fast-track protocol significantly decreased the number of complications (16 of 76 in the fast-track group vs 37 of 75 in the standard care group; P = .0014), resulting in shorter hospital stays (median, 5 days; range, 2-30 vs 9 days, respectively; range, 6-30; P < .0001). There was a trend toward less severe complications in the fast-track group. A multiple logistic regression analysis revealed fluid administration greater than the restriction limits (odds ratio, 4.198; 95% confidence interval, 1.7-10.366; P = .002) and a nonfunctioning epidural analgesia (odds ratio, 3.365; 95% confidence interval, 1.367-8.283; P = .008) as independent predictors of postoperative complications. CONCLUSIONS: The fast-track program reduces the rate of postoperative complications and length of hospital stay and should be considered as standard care. Fluid restriction and an effective epidural analgesia are the key factors that determine outcome of the fast-track program.

Paraoxonase2 polymorphisms are associated with nephropathy in Type II diabetes.

AIMS/HYPOTHESIS: Paraoxonase is a member of a multigene family of three genes. Paraoxonase2 gene polymorphisms have been associated with coronary heart disease in non-diabetic patients and with an increased fasting glycaemia in patients with Type II (non-insulin-dependent) diabetes mellitus. We tested the hypothesis of whether paraoxonase1 and paraoxonase2 polymorphisms were associated with diabetic nephropathy. METHODS: Our case-control study of 299 Swiss patients with Type II diabetes included 147 patients with confirmed diabetic nephropathy. RESULTS: In univariate analyses the two paraoxonase2 polymorphisms were associated with diabetic nephropathy. When subjected to multivariate analyses, both paraoxonase2 polymorphisms remained statistically associated with diabetic nephropathy independent of traditional risk factors (paraoxonase2-148: OR = 2.53, p = 0.003; paraoxonase2-311: OR = 2.67, p = 0.002). In addition, BMI interacted with paraoxonase2 polymorphisms as a risk factor of nephropathy. CONCLUSIONS/INTERPRETATION: The paraoxonase2 gene polymorphisms were significantly associated with diabetic nephropathy independent of traditional risk factors in Type II diabetic patients. The susceptibility to diabetic nephropathy was intensified by the degree of obesity. Pathophysiological pathways should be investigated and could be involved in insulin resistance or lipids metabolism or both.