UNDERSTANDING THE RELATIONSHIP BETWEEN THE BROWN MARMORATED STINK BUG, HALYOMORPHA HALYS (STÅL), AND ITS SYMBIONT, PANTOEA CARBEKII, WITH IMPLICATIONS FOR STINK BUG MANAGEMENT

Symbiotic relationships between insects and beneficial microbes are very common in nature, especially within the Hemiptera. The brown marmorated stink bug, Halyomorpha halys Stål, harbors a symbiont, Pantoea carbekii, within the fourth region of the midgut in specialized crypts. In this dissertation, I explored this insect- microbe relationship. I determined that the brown marmorated stink bug is heavily reliant on its symbiont, and that experimental removal of the symbiont from the egg mass surface prior to nymphal acquisition led to lower survival, longer development, lower fecundity, and aberrant nymphal behavior. Additionally, I determined that even when the symbiont is acquired and housed in the midgut crypts, it is susceptible to stressors. Stink bugs reared at a higher temperature showed lower survival, longer development, and a cease in egg mass production, and when bugs were screened for their symbiont, fewer had successfully retained it while under heat stress. Finally, with the knowledge that the stink bug suffers decreases in fitness when its symbiont is missing or stressed, I wanted to determine if targeting the symbiont was a possible management technique for the stink bug. I tested the efficacy of a number of different insecticidal and antimicrobial products to determine whether prevention of symbiont acquisition from the egg mass was possible, and results indicated that transmission of the symbiont from the egg mass to the newly hatched nymph was negatively impacted when certain products were applied (namely surfactants or products containing surfactants). Additionally, direct effects on hatch rate and survival were reported for certain products, namely the insect growth regulator azadirachtin, which suggests that nymphs can pick up residues from the egg mass surface while probing for the symbiont. I conclude that P. carbekii plays a critically important role in the survival of its host, the brown marmorated stink bug, and its presence on the egg mass surface before nymphal hatch makes it targetable as a potential management technique.

Thermotolerant cyclamen with reduced acrolein and methyl vinyl ketone

Reduced levels of trienoic fatty acids (TAs) in chloroplast membranes induce thermotolerance in several plant species, but the underlying mechanisms remain unclear. TA peroxidation in plant cell membranes generates cytotoxic, TA-derived compounds containing alpha,beta-unsaturated carbonyl groups. The relationship between low TA levels and the amounts of cytotoxic TA-derived compounds was examined using thermotolerant transgenic cyclamen (Cyclamen persicum Mill.) with low TA contents. Changes in the levels of the cytotoxic TA-derived acrolein (ACR), methyl vinyl ketone (MVK), (E)-2-hexenal, 4-hydroxy-2-nonenal, and malondialdehyde were analysed in the leaf tissues of wild-type (WT) and thermotolerant transgenic cyclamen under heat stress. Levels of ACR and MVK in the WT increased in parallel with the occurrence of heat-induced tissue damage, whereas no such changes were observed in the thermotolerant transgenic lines. Furthermore, exogenous ACR and MVK infiltrated into leaves to concentrations similar to those observed in heat-stressed WT leaves caused similar disease symptoms. These results suggest that thermotolerance in transgenic cyclamen depends on reduced production rates of ACR and MVK under heat stress, due to the low level of TAs in these plants.

Variações anátomo-fisiológicas face à aclimatação sazonal - estudo em vacas leiteiras com diferente potencial leiteiro

O presente estudo teve como objetivo a caracterização de variações anátomo-fisiológicas que decorrem do processo de aclimatação sazonal em bovinos leiteiros com alto (Altas) e baixo (Baixas) potencial leiteiro, com vista a selecionar animais que conciliem bons desempenhos produtivos e índices de adaptabilidade que possibilitem a mudança de homeostase face às condições climáticas presentes no clima mediterrânico. O estudo foi realizado numa herdade comercial, situada no Alentejo, utilizando 13 vacas multíparas (6 Baixas e 7 Altas), durante três períodos: P1 (animais aclimatados ao verão; presença de stresse térmico); P2 (animais aclimatados ao verão; termoneutralidade); P3 (animais aclimatados ao inverno; termoneutralidade). Em stresse térmico (P1), verificaram-se maiores esforços termolíticos e maiores armazenamentos de calor no grupo das Altas. Observou-se também que a produção de leite das Altas foi afetada pelo stresse térmico, evidênciando uma redução 24-48h após os valores de temperatura retal mais elevados. Nesta situação, as Baixas apresentaram uma variação na produção oposta à das Altas. Em P1, os valores de proteína e de gordura no leite foram significativamente mais baixos que em P3, em ambos os grupos. A ureia no leite foi significativamente mais elevada nas Altas durante o P1, revelando potencial como biomarcador de stresse térmico. Do P1 para o P3 obser-vou-se uma redução gradual do hematócrito, da hemoglobina e da triiodotironina (T3). As Altas apresentaram uma maior redução de triiodotironina (T3) que as Baixas, como consequência de uma maior intensidade de aclimatação. Nos pelos não se registaram diferenças entre os períodos, o que contrasta com alguma bibliografia. Porém, a ausência da insolação direta poderá ter sido um fator determinante; ABSTRACT: The main objective of the present study was the characterization of anatomical and physiological variations that occur in the seasonal acclimatization process of dairy cows with high (Altas) and low (Baixas) milk yield potential. In this way it should be possible to do a selection of animals with good productive traits and also with adaptability indexes that allow a change in homeostasis to cope with the climatic conditions of the mediterranean climate. Meteorological, clinical, productive, physiological and anatomical data were collected. The study was conducted in an Alentejo's dairy farm, using 13 multiparous cows (6 with low milk yield and 7 with high milk yield), during three periods: P1 (animals acclimated to summer, in heat stress); P2 (animals acclimated to summer, thermoneutrality); P3 (animals acclimated to winter; thermoneutrality). In thermal stress (P1), the high milk yield group (Altas) shown greater thermolytic efforts and also higher heat storage. The milk yield in this group was also affected by heat stress, showing a decrease in production when the rectal temperature increased, with a delay of 24-42 hours. In this situation the Baixas group showed an opposite milk production variation. In P1, the protein and fat milk content was lower than in P3, in both groups. Milk urea levels were significantly higher during P1 in the Altas group, revealing potential as an heat stress biomarker. Hematocrit, hemoglobin and triiodothyronine (T3) values gradualy decreased from P1 to P3. T3 values were lower in Altas than in Baixas group, as a consequence of a more intense acclimatization. The hair analysis didn’t show the standard seasonal acclimatization process, indicating the absence of direct solar radiation as a determinant factor.

Correlations between thermal environment and egg quality of two layer commercial strains

Heat stress limits the productivity of laying hens, as reflected by egg production and egg quality. The present study aimed at showing the correlations between egg quality parameters and environmental variables recorded on the day eggs were laid and on the previous days. Birds were housed in battery cages in a commercial poultry house. Main component analyses were used to verify associations between environmental and production variables, and Pearson's linear correlation tests were used to further investigate those associations. Analyses were carried out separately for to layer strains, Dekalb® White and Hy-Line® w36, and the variables egg weight (g), eggshell weight (g), specific gravity (g/cm³) and eggshell percentage (%) were compared with the environmental variables of the same day of the production, and one, two, three, and four days before egg production. Sound intensity measured inside the houses was positively associated with the quality parameters of eggs produced on the next day. Thermal environmental variables affected the egg quality differently in each strain, particularly air temperature, internal roof tile temperature, relative humidity, and air velocity. Ammonia concentration measured inside the houses was lower than 1ppm, and did not affect production performance.

Correlations among behavior, performance and environment in broiler breeders using multivariate analysis

Animal welfare issues have received much attention not only to supply farmed animal requirements, but also to ethical and cultural public concerns. Daily collected information, as well as the systematic follow-up of production stages, produces important statistical data for production assessment and control, as well as for improvement possibilities. In this scenario, this research study analyzed behavioral, production, and environmental data using Main Component Multivariable Analysis, which correlated observed behaviors, recorded using video cameras and electronic identification, with performance parameters of female broiler breeders. The aim was to start building a system to support decision-making in broiler breeder housing, based on bird behavioral parameters. Birds were housed in an environmental chamber, with three pens with different controlled environments. Bird sensitivity to environmental conditions were indicated by their behaviors, stressing the importance of behavioral observations for modern poultry management. A strong association between performance parameters and the behavior at the nest, suggesting that this behavior may be used to predict productivity. The behaviors of ruffling feathers, opening wings, preening, and at the drinker were negatively correlated with environmental temperature, suggesting that the increase of in the frequency of these behaviors indicate improvement of thermal welfare.

Estimating mortality in laying hens as the environmental temperature increases

Layer mortality due to heat stress is an important economic loss for the producer. The aim of this study was to determine the mortality pattern of layers reared in the region of Bastos, SP, Brazil, according to external environment and bird age. Data mining technique were used based on monthly mortality records of hens in production, 135 poultry houses, from January 2004 to August 2008. The external environment was characterized according maximum and minimum temperatures, obtained monthly at the meteorological station CATI in the city of Tupa, SP, Brazil. Mortality was classified as normal (<= 1.2%) or high (> 1.2%), considering the mortality limits mentioned in literature. Data mining technique produced a decision tree with nine levels and 23 leaves, with 62.6% of overall accuracy. The hit rate for the High class was 64.1% and 59.9% for Normal class. The decision tree allowed finding a pattern in the mortality data, generating a model for estimating mortality based on the thermal environment and bird age.

Studying the role of the strawberry Fra protein family in the flavonoid metabolism during fruit ripening

Strawberry fruits are highly appreciated worldwide due to their pleasant flavor and aroma and to the health benefits associated to their consumption. An important part of these properties is due to their content in secondary metabolites, especially phenolic compounds, of which flavonoids are the most abundant in the strawberry fruit. Although the flavonoid biosynthesis pathway is uncovered, little is known about its regulation. The strawberry Fra a (Fra) genes constitute a large family of homologs of the major birch pollen allergen Bet v 1 and for which no equivalents exist in Arabidopsis. Our group has shown that Fra proteins are involved in the formation of colored compounds in strawberries (Muñoz et al., 2010), which mainly depends on the production of certain flavonoids; that they are structurally homologs to the PYR/PYL/RCAR Arabidopsis ABA receptor, and that they are able to bind flavonoids (Casañal et al., 2013). With these previous results, our working hypothesis is that the Fra proteins are involved in the regulation of the flavonoids pathway. They would mechanistically act as the ABA receptor, binding a protein interactor and a ligand to regulate a signaling cascade and/or act as molecular carriers. The main objective of this research is to characterize the Fra family in strawberry and gain insight into their role in the flavonoid metabolism. By RNAseq expression analysis in ripening fruits we have identified transcripts for 10 members of the Fra family. Although expressed in all tissues analyzed, each family member presents a unique pattern of expression, which suggests functional specialization for each Fra protein. Then, our next approach was to identify the proteins that interact with Fras and their ligands to gain knowledge on the role that these proteins play in the flavonoids pathway. To identify the interacting partners of Fras we have performed a yeast two hybrid (Y2H) screening against cDNA libraries of strawberry fruits at the green and red stages. A protein that shares a 95% homology to the Heat stress transcription factor A-4-C like of Fragaria vesca (HSA4C) interacts specifically with Fra1 and not with other family members, which suggests functional diversification of Fra proteins in specific signaling pathways. The Y2H screening is not yet saturated, so characterization of other interacting proteins with other members of the Fra family will shed light on the functional diversity within this gene family. This research will contribute to gain knowledge on how the flavonoid pathway, and hence, the fruit ripening, is regulated in strawberry; an economically important crop but for which basic research is still very limited. References: Muñoz, C, et al. (2010). The Strawberry Fruit Fra a Allergen Functions in Flavonoid Biosynthesis. Molecular Plant, 3(1): 113–124. Casañal, A, et al (2013). The Strawberry Pathogenesis-related 10 (PR-10) Fra a Proteins Control Flavonoid Biosynthesis by Binding Metabolic Intermediates. Journal of Biological Chemistry, 288(49): 35322–35332.

Proteomic analysis of Rhizobium freirei PRF 81 responses to low pH.

Rhizobium freirei PRF 81 is employed in common bean commercial inoculants in Brazil, due to its outstanding efficiency in fixing nitrogen, competitiveness and tolerance to abiotic stresses. Among the environmental conditions faced by rhizobia in soils, acidity is perhaps the encountered most, especially in Brazil. So, we used proteomics based approaches to study the responses of PRF 81 to a low pH condition. R. freirei PRF 81 was grown in TY medium until exponential phase in two treatments: pH 6,8 and pH 4,8. Whole-cell proteins were extracted and separated by two-dimensional gel electrophoresis, using IPG-strips with pH range 4-7 and 12% polyacrilamide gels. The experiment was performed in triplicate. Protein spots were detected in the high-resolution digitized gel images and analyzed by Image Master 2D Platinum v 5.0 software. Relative volumes (%vol) of compared between the two conditions tested and were statistically evaluated (p ≤ 0.05). Even knowing that R. freirei PRF 81 can still grow in more acid conditions, pH 4.8 was chosen because didn´t affect significantly the bacterial growth kinetics, a factor that could compromise the analysis. Using a narrow pH range, the gel profiles displayed a better resolution and reprodutibility than using broader pH range. Spots were mostly concentrated between pH 5-7 and molecular masses between 17-95 kDa. From the six hundred well-defined spots analyzed, one hundred and sixty-three spots presented a significant change in % vol, indicating that the pH led to expressive changes in the proteome of R. freirei PRF 81. Of these, sixty-one were up-regulated and one hundred two was downregulated in pH 4.8 condition. Also, fourteen spots were only identified in the acid condition, while seven spots was exclusively detected in pH 6.8. Ninety-five differentially expressed spots and two exclusively detected in pH 4,8 were selected for Maldi-Tof identification. Together with the genome sequencing and the proteome analysis of heat stress, we will search for molecular determinants of PRF 81 related to capacity to adapt to stressful tropical conditions.

Tempo gasto em pastejo por novilhas Girolando em sistema integração Lavoura-Pecuária-Floresta.

Objetivou-se com esse trabalho avaliar o tempo gasto em pastejo por novilhas Girolando em área de integração Lavoura-Pecuária (iLP) e integração Lavoura-Pecuária-Floresta (iLPF). Oito novilhas ¾ Holandês x Gir com média de 25 meses de idade e 268 kg de peso vivo foram distribuídas em delineamento cross-over 2x2 (dois sistemas x dois períodos de avaliação) para coleta de dados de bioacústica por 48 horas utilizando gravadores MP3 fixados em cabrestos. Esses dados foram analisados pelo software Audacity®. As médias dos tempos (em minutos) gastos com a atividade de pastejo em cada tratamento (iLP e iLPF) nos períodos diurno e noturno foram comparadas pelo Teste de Tuckey a 5% de probabilidade. Os animais pastejaram mais tempo no período diurno em relação ao noturno (509,11 vs. 233,03 minutos, respectivamente). Houve diferença significativa entre os sistemas durante o período diurno, em que as novilhas no sistema iLPF pastejaram por maior tempo que as do iLP (581,35 vs. 433,88 minutos, respectivamente). Observou-se também que a atividade de pastejo ao longo de todo o dia foi maior no iLPF do que no iLP (333,44 vs. 409,71 minutos, respectivamente). Novilhas Girolando gastam mais tempo pastejando durante o dia e em pastagens com componente arbóreo.

The Effect of Climate Variability on Main Components of Cow Milk in Iran

The main purpose of this study is to assess the relationship between six bioclimatic indices for cattle (temperature humidity (THI), environmental stress (ESI), equivalent temperature (ESI), heat load (HLI), modified heat load (HLInew) and respiratory rate predictor(RRP)) and fundamental milk components (fat, protein, and milk yield) considering uncertainty. The climate parameters used to calculate the climate indices were taken from the NASA-Modern Era Retrospective-Analysis for Research and Applications (NASA-MERRA) reanalysis from 2002 to 2010. Cow milk data were considered for the same period from April to September when cows use natural pasture, with possibility for cows to choose to stay in the barn or to graze on the pasture in the pasturing system. The study is based on a linear regression analysis using correlations as a summarizing diagnostic. Bootstrapping is used to represent uncertainty estimation through resampling in the confidence intervals. To find the relationships between climate indices (THI, ETI, HLI, HLInew, ESI and RRP) and main components of cow milk (fat, protein and yield), multiple liner regression is applied. The least absolute shrinkage selection operator (LASSO) and the Akaike information criterion (AIC) techniques are applied to select the best model for milk predictands with the smallest number of climate predictors. Cross validation is used to avoid over-fitting. Based on results of investigation the effect of heat stress indices on milk compounds separately, we suggest the use of ESI and RRP in the summer and ESI in the spring. THI and HLInew are suggested for fat content and HLInew also is suggested for protein content in the spring season. The best linear models are found in spring between milk yield as predictands and THI, ESI,HLI, ETI and RRP as predictors with p-value < 0.001 and R2 0.50, 0.49. In summer, milk yield with independent variables of THI, ETI and ESI show the highest relation (p-value < 0.001) with R2 (0.69). For fat and protein the results are only marginal. It is strongly suggested that new and significant indices are needed to control critical heat stress conditions that consider more predictors of the effect of climate variability on animal products, such as sunshine duration, quality of pasture, the number of days of stress (NDS), the color of skin with attention to large black spots, and categorical predictors such as breed, welfare facility, and management system. This methodology is suggested for studies investigating the impacts of climate variability/change on food quality/security, animal science and agriculture using short term data considering uncertainty or data collection is expensive, difficult, or data with gaps.

Comportamiento metabólico, fisiológico y electrocardiográfico en trabajadores expuestos a altas temperaturas en una empresa líder en la explotación y coquización del carbón en Boyacá, Colombia, 2016

Objetivo: Establecer la relación entre la exposición ocupacional a altas temperaturas o sobrecarga térmica con el comportamiento fisiológico, metabólico y electrocardiográfico. Métodos: estudio de corte transversal, donde se incluyeron dos grupos (expuesto y no expuesto a altas temperaturas) en una empresa minera, en el departamento de Boyacá, Colombia, en el año 2016. El número de participantes fue de 160 trabajadores del género masculino, grupo expuesto (n=86) y grupo no expuesto (n=74). La exposición ocupacional a sobrecarga térmica se evaluó con el índice de temperatura de globo y bulbo húmedo (TGBH), el comportamiento fisiológico con el índice de costo cardiaco relativo (ICCR) con mediciones de frecuencia cardiaca (FC), el comportamiento metabólico con la determinación del colesterol total (CT), colesterol de alta densidad (C-HDL), colesterol de baja densidad (C-LDL), triglicéridos (TG) y glicemia basal (GL). Las alteraciones electrocardiográficas con la toma de Electrocardiograma de 12 derivaciones. También fueron evaluadas variables antropométricas, tensión arterial, hábitos y antecedentes de enfermedad cardiovascular en ambos grupos. Resultados: incrementos significativos del ICCR (p<0.001) y la carga física (p<0.001) fueron encontrados en los trabajadores expuestos a altas temperaturas. Los índices lipídicos y glicemia, así como los antecedentes personales cardiovasculares, IMC, consumo de cigarrillo y consumo de alcohol, no mostraron significancia. El antecedente familiar de ACV (p=0.043) y el EKG alterado (p=0.011) mostraron una asociación significativa con la exposición a altas temperaturas. El modelo de regresión lineal múltiple explicó la relación entre el incremento del ICCR y la exposición a altas temperaturas (β=4,213, IC 95%: 1.57,6.85) ajustado por variables fisiológicas y electrocardiográficas. Conclusiones: La exposición ocupacional a altas temperaturas, presenta asociación con las alteraciones cardiovasculares a nivel fisiológico y electrocardiográfico, aumentando el ICCR y la carga física de trabajo (GE trabajo).

Thermoregulatory response in hair sheep and shorn wool sheep

This study’s main goal was to evaluate the thermoregulatory responses velocity through the variation of rectal temperature (RT), related to the thermolytic pathways, respiratory rate (RR) and sweating rate (SR) among different sheep breeds. Ninety female sheep, eighteen of each breed: Santa Ines and Morada Nova (Brazilian hair breeds), Texel, Suffolk and Ile de France (wool breeds) were challenged during three non-consecutive summer days (22◦42′S, 47◦18′W, and 570m of altitude, maximum air temperature of 33.5◦C, average relative humidity of 52±6.9%). The physiological variables were registered at 0800h (T1), 1300 h (T2: after 2 h of shade rest), 1400 h (T3) (after one hour of sun exposure) and in the shade at 1415 h (T4), 1430 h (T5), 1445 h (T6) and 1500 h (T7) and a thermotolerance index (TCI) was calculated as (10-(T7 to T4)-T1). The statistical analysis was performed by a mathematical model including the fixed effects of breeds and time frames, and the interaction between these effects, besides random effects such as animal and day. The Santa Ines breed presented the lowest RT after sun exposure (39.3 ± 0.12 ◦ C; P < 0.05) and it was the only one to recover morning RT 60 min after heat stress (38.7 and 38.9 for 1300 h and 1500 h; P > 0.05). Hair breeds presented RR lower (P < 0.05) than wool breeds. Although thick wool or hair thickness differs among and within hair and wool breeds (P < 0.05), SR did not differ among breeds and time (227.7 ± 16.44 g m−2 h−1 ; P > 0.05). The thermotolerance index did not differ among breeds, but it showed similar response (P > 0.05) 45 min or 1 h of shade after sun exposure. One week post shearing is not enough to wool breeds present to show thermotolerance similar to hair breeds.

Water stress induced alterations in ornithine aminotransferase of ragi (Eleusine coracana): Protection by proline against heat inactivation and denaturation by urea and guanidinium chloride

Water stress resulted in a specific response leading to a large and significant increase (80-fold) in free proline content of ragi (Eleusine coracana) leaves and seedlings. L-Proline protected ornithine aminotransferase, an enzyme in the pathway for proline biosynthesis, isolated from normal and stressed ragi leaves against heat inactivation and denaturation by urea and guanidinium chloride. The protection of the stressed enzyme by L-proline was much more complete than that of the enzyme isolated from normal leaves. While L-ornithine, one of the substrates, protected the stressed enzyme against inactivation, it enhanced the rate of inactivation of the normal enzyme. α-Ketoglutarate protected both the normal and stressed enzyme against inactivation and denaturation. These results support the suggestion that ornithine aminotransferase has undergone a structural alteration during water stress. In view of the causal relationship between elevated temperature and water stress of plants under natural conditions, the protection afforded by proline against inactivation and denaturation of the enzyme from stressed leaves assumes significance. These results provide an explanation for a possible functional importance of proline accumulation during water stress.

Arabidopsis thaliana J-class heat shock proteins: cellular stress sensors

Plants are sessile organisms that have evolved a variety of mechanisms to maintain their cellular homeostasis under stressful environmental conditions. Survival of plants under abiotic stress conditions requires specialized group of heat shock protein machinery, belonging to Hsp70:J-protein family. These heat shock proteins are most ubiquitous types of chaperone machineries involved in diverse cellular processes including protein folding, translocation across cell membranes, and protein degradation. They play a crucial role in maintaining the protein homeostasis by reestablishing functional native conformations under environmental stress conditions, thus providing protection to the cell. J-proteins are co-chaperones of Hsp70 machine, which play a critical role by stimulating Hsp70s ATPase activity, thereby stabilizing its interaction with client proteins. Using genome-wide analysis of Arabidopsis thaliana, here we have outlined identification and systematic classification of J-protein co-chaperones which are key regulators of Hsp70s function. In comparison with Saccharomyces cerevisiae model system, a comprehensive domain structural organization, cellular localization, and functional diversity of A. thaliana J-proteins have also been summarized. Electronic supplementary material The online version of this article (doi:10.1007/s10142-009-0132-0) contains supplementary material, which is available to authorized users.