SS.PP. Toletanorum quotquot extant Opera : tomus primus [-tertius]

Nunc primum simul edita, ad codices mss. recognita, nonnullis notis illustrata atque in duos tomos distributa.

Evaluation of microbial adjuncts and their effect on the ripening of cheddar cheese

A bacteriocin-producing strain of Lactobacillus paracasei DPC 4715 was used as an adjunct culture in Cheddar cheese in order to control the growth of “wild” nonstarter lactic acid bacteria. No suppression of growth of the indicator strain was observed in the experimental cheese. The bacteriocin produced by Lactobacillus paracasei DPC 4715 was sensitive to chymosin and cathepsin D and it may have been cleaved by the rennet used for the cheese manufactured or by indigenous milk proteases. A series of studies were performed using various microbial adjuncts to influence cheese ripening. Microbacterium casei DPC 5281, Corynebacterium casei DPC 5293 and Corynebacterium variabile DPC 5305 were added to the cheesemilk at level of 109 cfu/ml resulting in a final concentration of 108 cfu/g in Cheddar cheese. The strains significantly increased the level of pH 4.6-soluble nitrogen, total free amino acids after 60 and 180 d of ripening and some individual free amino acids after 180 d. Yarrowia lipolytica DPC 6266, Yarrowia lipolytica DPC 6268 and Candida intermedia DPC 6271 were used to accelerate the ripening of Cheddar cheese. Strains were grown in YG broth to a final concentration of 107 cfu/ml, microfluidized, freeze-dried and added to the curd during salting at level of 2% w/w. The yeasts positively affected the primary, secondary proteolysis and lipolysis of cheeses and had aminopeptidase, dipeptidase, esterase and 5’ phosphodiestere activities that contributed to accelerate the ripening and improve the flavor of cheese. Hafia alvei was added to Cheddar cheesemilk at levels of 107 cfu/ml and 108 cfu/ml and its contribution during ripening was evaluated. The strain significantly increased the level of pH 4.6-soluble nitrogen, total free amino-acids, and some individual free amino-acids of Cheddar cheese, whereas no differences in the urea-polyacrylamide gel electrophoresis (urea-PAGE) electrophoretograms of the cheeses were detected. Hafia alvei also significantly increased the level of some biogenic amines. A low-fat Cheddar cheese was made with Bifidobacterium animalis subsp. lactis, strain BB-12® at level of 108 cfu/ml, as a probiotic adjunct culture and Hi-Maize® 260 (resistant high amylose maize starch) at level of 2% and 4% w/v, as a prebiotic fiber which also played the role of fat replacer. Bifidobacterium BB-12 decreased by 1 log cycle after 60 d of ripening and remained steady at level of ~107 cfu/g during ripening. The Young’s modulus also increased proportionally with increasing levels of Hi-maize. Hencky strain at fracture decreased over ripening and increased with increasing in fat replacer. A cheese based medium (CBM) was developed with the purpose of mimicking the cheese environment at an early ripening stage. The strains grown in CBM showed aminopeptidase activity against Gly-, Arg-, Pro- and Phe-p-nitroanalide, whereas, when grown in MRS they were active against all the substrates tested. Both Lb. danicus strains grown in MRS and in CBM had aminotransferase activity towards aromatic amino acids (Phe and Trp) and also branched-chain amino acids (Leu and Val). Esterase activity was expressed against p-nitrophenyl-acetate (C2), pnitrophenyl- butyrate (C4) and p-nitrophenyl-palmitate (C16) and was significantly higher in CBM than in MRS.

Heterogeneous molybdate catalysts for the generation of singlet molecular oxygen (1Δg) from H2O2

The immobilisation of molybdate on Mg,Al-LDH leads to an active, heterogeneous catalyst that generates singlet molecular oxygen from hydrogen peroxide in the absence of soluble base

Immunization with cocktail of HIV-derived peptides in montanide ISA-51 is immunogenic, but causes sterile abscesses and unacceptable reactogenicity.

BACKGROUND: A peptide vaccine was produced containing B and T cell epitopes from the V3 and C4 Envelope domains of 4 subtype B HIV-1 isolates (MN, RF, CanO, & Ev91). The peptide mixture was formulated as an emulsion in incomplete Freund's adjuvant (IFA). METHODS: Low-risk, healthy adult subjects were enrolled in a randomized, placebo-controlled dose-escalation study, and selected using criteria specifying that 50% in each study group would be HLA-B7+. Immunizations were scheduled at 0, 1, and 6 months using a total peptide dose of 1 or 4 mg. Adaptive immune responses in16 vaccine recipients and two placebo recipients after the 2nd immunization were evaluated using neutralization assays of sera, as well as ELISpot and ICS assays of cryopreserved PBMCs to assess CD4 and CD8 T-cell responses. In addition, (51)Cr release assays were performed on fresh PBMCs following 14-day stimulation with individual vaccine peptide antigens. RESULTS: 24 subjects were enrolled; 18 completed 2 injections. The study was prematurely terminated because 4 vaccinees developed prolonged pain and sterile abscess formation at the injection site-2 after dose 1, and 2 after dose 2. Two other subjects experienced severe systemic reactions consisting of headache, chills, nausea, and myalgia. Both reactions occurred after the second 4 mg dose. The immunogenicity assessments showed that 6/8 vaccinees at each dose level had detectable MN-specific neutralizing (NT) activity, and 2/7 HLA-B7+ vaccinees had classical CD8 CTL activity detected. However, using both ELISpot and ICS, 8/16 vaccinees (5/7 HLA-B7+) and 0/2 controls had detectable vaccine-specific CD8 T-cell responses. Subjects with moderate or severe systemic or local reactions tended to have more frequent T cell responses and higher antibody responses than those with mild or no reactions. CONCLUSIONS: The severity of local responses related to the formulation of these four peptides in IFA is clinically unacceptable for continued development. Both HIV-specific antibody and T cell responses were induced and the magnitude of response correlated with the severity of local and systemic reactions. If potent adjuvants are necessary for subunit vaccines to induce broad and durable immune responses, careful, incremental clinical evaluation is warranted to minimize the risk of adverse events. TRIAL REGISTRATION: ClinicalTrials.gov NCT00000886.

Structure, function, and phylogeny of the mating locus in the Rhizopus oryzae complex.

The Rhizopus oryzae species complex is a group of zygomycete fungi that are common, cosmopolitan saprotrophs. Some strains are used beneficially for production of Asian fermented foods but they can also act as opportunistic human pathogens. Although R. oryzae reportedly has a heterothallic (+/-) mating system, most strains have not been observed to undergo sexual reproduction and the genetic structure of its mating locus has not been characterized. Here we report on the mating behavior and genetic structure of the mating locus for 54 isolates of the R. oryzae complex. All 54 strains have a mating locus similar in overall organization to Phycomyces blakesleeanus and Mucor circinelloides (Mucoromycotina, Zygomycota). In all of these fungi, the minus (-) allele features the SexM high mobility group (HMG) gene flanked by an RNA helicase gene and a TP transporter gene (TPT). Within the R. oryzae complex, the plus (+) mating allele includes an inserted region that codes for a BTB/POZ domain gene and the SexP HMG gene. Phylogenetic analyses of multiple genes, including the mating loci (HMG, TPT, RNA helicase), ITS1-5.8S-ITS2 rDNA, RPB2, and LDH genes, identified two distinct groups of strains. These correspond to previously described sibling species R. oryzae sensu stricto and R. delemar. Within each species, discordant gene phylogenies among multiple loci suggest an outcrossing population structure. The hypothesis of random-mating is also supported by a 50:50 ratio of plus and minus mating types in both cryptic species. When crossed with tester strains of the opposite mating type, most isolates of R. delemar failed to produce zygospores, while isolates of R. oryzae produced sterile zygospores. In spite of the reluctance of most strains to mate in vitro, the conserved sex locus structure and evidence for outcrossing suggest that a normal sexual cycle occurs in both species.

Expansion of the alpha 2-adrenergic receptor family: cloning and characterization of a human alpha 2-adrenergic receptor subtype, the gene for which is located on chromosome 2.

Pharmacologic, biochemical, and genetic analyses have demonstrated the existence of multiple alpha 2-adrenergic receptor (alpha 2AR) subtypes. We have cloned a human alpha 2AR by using the polymerase chain reaction with oligonucleotide primers homologous to conserved regions of the previously cloned alpha 2ARs, the genes for which are located on human chromosomes 4 (C4) and 10 (C10). The deduced amino acid sequence encodes a protein of 450 amino acids whose putative topology is similar to that of the family of guanine nucleotide-binding protein-coupled receptors, but whose structure most closely resembles that of the alpha 2ARs. Competition curve analysis of the binding properties of the receptor expressed in COS-7 cells with a variety of adrenergic ligands demonstrates a unique alpha 2AR pharmacology. Hybridization with somatic cell hybrids shows that the gene for this receptor is located on chromosome 2. Northern blot analysis of various rat tissues shows expression in liver and kidney. The unique pharmacology and tissue localization of this receptor suggest that this is an alpha 2AR subtype not previously identified by classical pharmacological or ligand binding approaches.

Infection of monkeys by simian-human immunodeficiency viruses with transmitted/founder clade C HIV-1 envelopes.

Simian-human immunodeficiency viruses (SHIVs) that mirror natural transmitted/founder (T/F) viruses in man are needed for evaluation of HIV-1 vaccine candidates in nonhuman primates. Currently available SHIVs contain HIV-1 env genes from chronically-infected individuals and do not reflect the characteristics of biologically relevant HIV-1 strains that mediate human transmission. We chose to develop clade C SHIVs, as clade C is the major infecting subtype of HIV-1 in the world. We constructed 10 clade C SHIVs expressing Env proteins from T/F viruses. Three of these ten clade C SHIVs (SHIV KB9 C3, SHIV KB9 C4 and SHIV KB9 C5) replicated in naïve rhesus monkeys. These three SHIVs are mucosally transmissible and are neutralized by sCD4 and several HIV-1 broadly neutralizing antibodies. However, like natural T/F viruses, they exhibit low Env reactivity and a Tier 2 neutralization sensitivity. Of note, none of the clade C T/F SHIVs elicited detectable autologous neutralizing antibodies in the infected monkeys, even though antibodies that neutralized a heterologous Tier 1 HIV-1 were generated. Challenge with these three new clade C SHIVs will provide biologically relevant tests for vaccine protection in rhesus macaques.

PDLA a potential new potent topical analgesic: a case report.

Polymer D-lactic acid (PDLA) is a hydrogel that has been shown to sequester L-lactate (lactate). This reaction is rapid, spontaneous, and non-enzymatic. Lactate has been shown to have many functions within the nervous system including its use as a secondary fuel to sustain neural activity and as a neuromodulator. In the central nervous system, lactate is produced in glial cells and shuttled to neurons to be used mostly as a fuel. Lactate dehydrogenase (LDH)1 is the predominant LDH isoform within neurons and unlike LDH5, it preferentially converts lactate to pyruvate which can be used to produce adenosine triphosphate (ATP). Considering that lactate is intimately involved in the sustenance of neural activity, PDLA was applied to an open wound and its effects were examined. The results showed that the application of PDLA induced topical analgesia. This may be the first report to demonstrate that sequestering lactate, a source of energy required to sustain the firing of action potentials in neurons, may produce analgesia.

Stereocomplexes Formed From Select Oligomers of Polymer d-lactic Acid (PDLA) and l-lactate May Inhibit Growth of Cancer Cells and Help Diagnose Aggressive Cancers-Applications of the Warburg Effect.

It is proposed that select oligomers of polymer d-lactic acid (PDLA) will form a stereocomplex with l-lactate in vivo, producing lactate deficiency in tumor cells. Those cancer cells that utilize transport of lactate to maintain electrical neutrality may cease to multiply or die because of lactate trapping, and those cancer cells that benefit from utilization of extracellular lactate may be impaired. Intracellular trapping of lactate produces a different physiology than inhibition of LDH because the cell loses the option of shuttling pyruvate to an alternative pathway to produce an anion. Conjugated with stains or fluorescent probes, PDLA oligomers may be an agent for the diagnosis of tissue lactate and possibly cell differentiation in biopsy specimens. Preliminary experimental evidence is presented confirming that PDLA in high concentrations is cytotoxic and that l-lactate forms a presumed stereocomplex with PDLA. Future work should be directed at isolation of biologically active oligomers of PDLA.

The accuracy and completeness for receipt of colorectal cancer care using Veterans Health Administration administrative data.

BACKGROUND: The National Comprehensive Cancer Network and the American Society of Clinical Oncology have established guidelines for the treatment and surveillance of colorectal cancer (CRC), respectively. Considering these guidelines, an accurate and efficient method is needed to measure receipt of care. METHODS: The accuracy and completeness of Veterans Health Administration (VA) administrative data were assessed by comparing them with data manually abstracted during the Colorectal Cancer Care Collaborative (C4) quality improvement initiative for 618 patients with stage I-III CRC. RESULTS: The VA administrative data contained gender, marital, and birth information for all patients but race information was missing for 62.1% of patients. The percent agreement for demographic variables ranged from 98.1-100%. The kappa statistic for receipt of treatments ranged from 0.21 to 0.60 and there was a 96.9% agreement for the date of surgical resection. The percentage of post-diagnosis surveillance events in C4 also in VA administrative data were 76.0% for colonoscopy, 84.6% for physician visit, and 26.3% for carcinoembryonic antigen (CEA) test. CONCLUSIONS: VA administrative data are accurate and complete for non-race demographic variables, receipt of CRC treatment, colonoscopy, and physician visits; but alternative data sources may be necessary to capture patient race and receipt of CEA tests.

Estimación satelital de la productividad primaria neta aérea de la vegetación herbácea del Caldenal

En la Región del Caldenal es necesario generar cambios en la implementación de prácticas de manejo de la ganadería, desarrollar una actividad sustentable y, a su vez, evitar el mayor deterioro del ecosistema y sus servicios. Dichos cambios deberían incluir el uso racional del la vegetación herbácea, combinado con pasturas C4, actividad que en los últimos años se expandió en la región. Para lograr la complementariedad de dichos recursos y poder estructurar estrategias de manejo, debe conocerse su funcionamiento, estacionalidad y productividad. La información satelital provee la ventaja de otorgar gran cantidad y calidad de datos, en distintas escalas espaciales y temporales. El presente trabajo explora algunas posibilidades para la aplicación de dichas herramientas en la Región del Caldenal. La finalidad es proponer estrategias para conocer en forma sencilla la productividad primaria neta aérea del sistema, como determinante de la receptividad animal y el funcionamiento del ecosistema. Se utilizaron imágenes correspondientes a 60 potreros, distribuidos en seis localidades de la Región del Caldenal, Provincia de La Pampa, con bosque de caldén ("fachinal"), sabanas ("Caldenal"), pasturas de Eragrostis curvula y de Panicum coloratum. Con dichas imágenes se evaluó la variabilidad del Índice de Vegetación Normalizado (IVN) en el tiempo, relacionándolo con el funcionamiento ecosistémico (Capítulo 2), se propuso una metodología para discriminar el IVN de la vegetación herbácea y leñosa en los potreros de sabana (Capítulo 3) y se estimó la Productividad Primaria Neta Aérea de los distintos recursos, realizando comparaciones de estacionalidad y producción, y correlaciones con variables ambientales (Capítulo 4). El presente trabajo demuestra las potencialidades de uso de herramientas satelitales para estimar diferentes atributos de los recursos naturales y cultivados del Caldenal. El ajuste de dichas técnicas en la región resulta clave para el diseño de estrategias de manejo ganadero sustentables y conservación de los recursos.

Efectos de la relación entre hongos micorrícicos arbusculares y gramíneas forrajeras bajo defoliación y distinto suministro de fósforo

La asociación entre raíces de plantas y hongos micorrícicos arbusculares (HMA) incrementa la absorción de fósforo. Sin embargo, la defoliación puede influenciar el balance de la simbiosis y la recuperación de las plantas. El objetivo de esta tesis fue estudiar el efecto de la simbiosis micorrícica y el suministro de fósforo, sobre el crecimiento y la recuperación post-defoliación de gramíneas forrajeras cultivadas. Para esto, en primer lugar, se caracterizó la respuesta micorrícica de seis especies de gramíneas (tres templadas (C3) : i) Dactylis glomerata, ii) Elymus elongatus y iii) Schedonorus arundinaceus; y tres tropicales (C4) : iv) Paspalum dilatatum, v) Panicum coloratum y vi) Urochloa brizantha) creciendo en déficit nutricional. En estas condiciones, las gramíneas tropicales presentaron mayor respuesta micorrícica que las templadas. En segundo lugar, se estudió el rol de las micorrizas y el suministro de fósforo sobre el crecimiento y la nutrición de E. elongatus y U. brizantha, dos especies con respuestas micorrícicas contrastantes. Al aumentar el suministro de fósforo, en E. elongatus (especie con menor respuesta micorrícic) ) y U. brizantha (especie con mayor respuesta micorrícica) el cambio en la producción de biomasa, entre plantas inoculadas y no-inoculadas, disminuyó. Sin embargo, mostraron diferencias en la eficiencia en el uso del fósforo: U. brizantha hizo un mayor aprovechamiento del fósforo que E. elongatus. Por último, se caracterizaron respuestas de recuperación post-defoliación bajo distinto suministro de fósforo. En E. elongatus en alto fósforo, hubo un efecto parasítico de la micorriza sobre la recuperación post-defoliación. Por el contrario, en U. brizantha no se detectó un compromiso entre defoliación y micorrizas. La información generada en esta tesis servirá de base para mejorar el conocimiento de aspectos básicos de la relación entre hongos micorrícicos arbusculares y gramíneas forrajeras, como también, para establecer pautas de conservación de las relaciones simbióticas en sistemas pastoriles.

Modificaciones en la vegetación y el suelo inducidos por el manejo del pastoreo en la estepa de halófitas de la pampa deprimida

La actividad agrícola impuso cambios en el uso de la tierra que condujo a un incremento en la concentración de animales en los ambientes marginales de la Pampa Deprimida entre los que se destaca la estepa de halófitas. En ese marco resulta primordial entender mejor los efectos del pastoreo sobre estos ambientes frágiles y pocos estudiados, para proponer herramientas de manejo en situaciones muy deterioradas por el pastoreo continuo. Esta tesis avanzó en el conocimiento de los cambios en la estructura de la vegetación y en las propiedades físicas y químicas del suelo de la estepa de halófitas como resultado de cambiar el manejo de pastoreo continuo por pastoreo rotativo o exclusión por varios años. La implementación del pastoreo rotativo o la exclusión aumentaron la riqueza y la diversidad florística, la cobertura total de broza y la de los grupos funcionales de gramíneas C3 anuales, C4 perennes, monocotiledoneas y leguminosas, y produjeron un contundente cambio en la composición florística. Los cambios florísticos dieron lugar a un aumento importante del valor forrajero de la comunidad vegetal. La metodología de pastoreo propuesta aumentó el contenido de materia orgánica y causaron descensos significativos en la salinidad y el pH, con respecto a las áreas bajo pastoreo continuo. En consonancia con ello, se produjeron mejoras en las propiedades físicas del suelo, como la disminución de la inestabilidad estructural y de la densidad aparente. Los resultados sugieren que la restauración de la estepa de halófitas es factible cuando se implementa el pastoreo rotativo o la exclusión después de ser pastoreadas continuamente.

Comportamiento de varias cepas de Azospirillum sobre trigo (Triticum aestivum) y sorgo (Sorghum vulgare)

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