Development of a multiplex polymerase chain reaction to detect five common Gram-negative bacteria of aquatic animals

A multiplex polymerase chain reaction (m-PCR) technique was developed as a rapid and accurate diagnostic tool for identifying five major Gram-negative bacilli -Vibrio vulnificus, V. parahaemolyticus, Aeromonas hydrophila, Chryseobacterium meningosepticum and Edwardsiella tarda- that cause major diseases in cultured aquatic animals in Taiwan. The expected amplicons for V. vulnificus, V. parahaemolyticus, A. hydrophila, C. meningosepticum and E. tarda were 410, 368, 685, 180 and 230bp, respectively. The assay was shown to be specific for the target pathogens. The sensitivities of detection were estimated to be 20.5fg～200pg of genomic DNA or 10(2) ～10(4) colony-forming units (cfu) of bacterial isolates when adopted as PCR templates. The m-PCR was capable of simultaneously amplifying target fragments from bacterial genome DNA mixed with the DNA extracted from viscera and tissues taken from fish without affecting the performance of the method.     

Histopathologic changes of the ear in canine models of mucopolysaccharidosis types I and VII

Mucopolysaccharidosis (MPS) types I and VII are inborn errors of metabolism caused by mutation of enzymes involved in glycosaminoglycan catabolism, which leads to intralysosomal accumulation of glycosaminoglycans. In children, severe forms of MPS I and VII are characterized by somatic and neurologic manifestations, including a poorly understood hearing loss. The purpose of this study is to describe the age-related histopathologic changes of the ear in spontaneous canine models of MPS I and VII. Pathologic changes in the ear were assessed in MPS I and VII dogs ranging from 1.6 to 9.3 months of age. Paraffin-embedded sections of the whole ear and Epon-embedded semithin sections of the cochlea were examined. The following lesions were blindly scored in the middle and inner ear: inflammation, cells vacuolization, thickening of osseous and membranous structures, perivascular vacuolated macrophages infiltration, and bone resorption. All dogs had lysosomal storage within cells of tympanic membrane, ossicles, tympanic bone and mucosa, cochlear bone, spiral ligament, limbus, and stria vascularis. The MPS I dogs mainly had progressive cochlear lesions. The MPS VII dogs had severe and early middle ear lesions, including chronic otitis media and bone resorption. The MPS I dog only partially recapitulates the pathology seen in humans; specifically, the dog model lacks inflammatory middle ear disease. In contrast, the MPS VII dog has severe inflammatory middle ear disease similar to that reported in the human. In conclusion, the canine MPS VII model appears to be a good model to study MPS VII-related deafness.     

Horse lumbrical muscle: possible structural and functional reorganization in regressive muscle

An anatomical study of horse lumbrical muscle (Lm) was carried out by light and electron microscopy in combination with immunochemical and cytochemical methods. Paraffin sections were subjected to haematoxylin and eosin (H & E) and Masson's trichrome staining for morphometric analysis. Paraffin sections were also used for immunostaining by anti-PGP 9.5 for reaction with nerve-protein associated-structures, anti-heat-shock protein 70 (hsp 70) for detection of gene expression changes, anti-fast myosin for the determination of muscle fibre types, and for detection of apoptotic gene expression of muscle fibres by the TUNEL method. The distribution of muscle fibre types on frozen sections was also examined by assaying ATPase (pH 4.2). We found that the proximal end of the tendon of the unipennate-shaped Lm binds to the deep digital flexor tendon, and the distal end of the Lm tendon connects to the medial surface of the palmar annular ligament. The Lm was not always present, but when found it varied in length greatly, up to 8 cm (muscle part alone), and weighed less than 1 g. The Lm was white, pale, or reddish in colour depending on the ratio of muscle fibre and connective tissue contents. The semi-tendinized regressive Lm was composed of rich vasculature, peripheral nerves, and nerve-like organs similar to the neuromuscular spindle (NMS). The extrafusal muscle fibres (e-lm) that surround the NMS were replaced with a thick outer capsule of connective tissues (CT) in the Lm nerve-like organ, which we named the neurotendinous capsule (NTC) organ. NTC organs exist alone or as multiple structures (up to eight) surrounded by a common outer capsule at the outermost CT ring. The NTC possesses several intrafusal muscle fibres (ifm) just as the NMS does. That the ifm was associated with nerve endings was confirmed by anti-PGP 9.5 and electron microscopic observation. Some muscle fibres in ifm and e-lm reacted with anti-fast twitch myosin and with anti-hsp 70. The e-lm exhibited at least two fibre types, determined by ATPase (pH 4.2) assay. The ifm exhibited mainly type I (slow twitch) fibres. No apoptotic gene expression was detected in either ifm or e-lm, suggesting the Lm is a vital organ. The degenerating fibres observed in ifm and e-lm indicate that the turnover rate of cytoplasmic components is accelerated. We attribute this phenomenon to the necessity for adaptation to new environmental demands. The surprising finding of tubular aggregates (TAs) in ifm of the NTC organ suggests that the Lm is continuously adapting. Some results related to variation in diameter of the collagen fibrils, isolation of the NTC organ and the myofibrillar protein constituents are also discussed. In conclusion, the so-called regressive Lm has rich vasculature, many peripheral nerves, and newly described NTC organs. The induction of heat-shock protein, lack of apoptotic gene expression in ifm and e-lm fibres, and TA formation in ifm suggest that horse Lm responds to environmental stress through reorganization and/or remodelling of cell constituents. We hypothesize that the horse Lm has lost its original role as a contractile element and changed to another function, likely as a vital nerve organ.     

Efficacy and cardiorespiratory effects of lumbosacral intrathecal 2% procaine and 2% xylazine with or without sedation in calves undergoing umbilical surgery

ABSTRACT

Aims: To compare the effects of intrathecal anaesthesia using procaine and xylazine, with and without sedation with I/V xylazine and butorphanol, on sedation and cardiorespiratory measures in calves undergoing umbilical surgery.

Methods: Male dairy calves, aged <3 months, were recruited that had enlargement of the umbilical stalk which was abnormal when palpated. They were assigned to receive either intrathecal injection between the sixth lumbar and first sacral vertebrae of 4?mg/kg of 2% procaine and 0.2?mg/kg 2% xylazine, with I/V injection of 0.02?mg/kg xylazine and 0.1?mg/kg of butorphanol (IT?+?SED group; n?=?6), or the same intrathecal injection and I/V injection of 0.9% saline (IT group; n?=?7). Surgery to correct abnormalities was carried out with calves positioned in dorsal recumbency. Rescue analgesia with injections of 2% procaine around the surgical wound was administered when movements triggered by surgery were observed. Post-operative analgesia was provided using I/V 0.5?mg/kg meloxicam. Duration of surgery was recorded, as well as degree of sedation, heart rate, systolic (SAP), diastolic (DAP) and mean (MAP) arterial blood pressure during surgery.

Results: All anaesthetic and surgical procedures were successfully performed. Mean total duration of surgery was similar for the IT?+?SED and the IT groups (30.33 (SD 10.09) and 31.00 (SD 10.21) minutes, respectively) (p?=?0.92). All calves were at least mildly sedated from 5 minutes after injections to the end of the surgery. One calf in the IT?+?SED group and three calves in the IT group required rescue analgesia when the umbilicus was manipulated. Between 0 and 10–15 minutes after injection, decreases in mean heart rate, SAP, MAP and DAP were observed in both groups. Mean SAP was lower in the IT?+?SED than the IT group. Hypotension (MAP<60?mm Hg) was present in four calves from the IT?+?SED group and in one from the IT group.

Conclusions and clinical relevance: Intrathecal administration of 2% procaine and 2% xylazine allowed the successful completion of umbilical surgery, but 30% of calves needed rescue analgesia during surgery. Clinically, the addition of I/V sedation seemed to provide better analgesia than intrathecal block alone but resulted in greater hypotension.     

The impact of a school-based nutrition education intervention on dietary intake and cognitive and attitudinal variables relating to fruits and vegetables

OBJECTIVE: To assess the impact of a school-based nutrition education intervention aimed at increasing the consumption of fruits and vegetables. DESIGN: The intervention programme increased the provision of fruits and vegetables in schools and provided a range of point-of-purchase marketing materials, newsletters for children and parents, and teacher information. Curriculum materials at age 6-7 and 10-11 years were also developed and utilised. Evaluation was undertaken with groups of younger (aged 6-7 years) and older (aged 10-11 years) children. Methods included 3-day dietary records with interview and cognitive and attitudinal measures at baseline, with follow-up at 9 months, in intervention and control schools. SETTING: The work was undertaken in primary schools in Dundee, Scotland. SUBJECTS: Subjects comprised 511 children in two intervention schools with a further 464 children from two schools acting as controls. RESULTS: Children (n=64) in the intervention schools had an average increase in fruit intake (133+/-1.9 to 183+/-17.0 g day(-1)) that was significantly (P<0.05) greater than the increase (100+/-11.7 to 107+/-14.2 g day(-1)) estimated in children (n=65) in control schools. No other changes in food or nutrient intake were detected. Increases in scores for variables relating to knowledge about fruits and vegetables and subjective norms were also greater in the intervention than in the control group, although taste preferences for fruits and vegetables were unchanged. CONCLUSIONS: It is concluded that a whole school approach to increasing intakes of fruits and vegetables has a modest but significant effect on cognitive and attitudinal variables and on fruit intake.     

Leisure-time and occupational physical activity in relation to cardiovascular risk factors and eating habits in French adults

OBJECTIVE: To investigate the relationships of two main physical activity domains (during leisure and at work) with cardiovascular risk factors and eating habits. DESIGN: Cross-sectional study. SETTING: Preventive medicine centre. SUBJECTS: In 5478 adults (32% women, aged 20-80 years) who consecutively underwent a standardised health examination, leisure-time physical activity (LTPA; i.e. non-sport leisure and sport activities), occupational physical activity (OPA) and eating habits were assessed by self-administered questionnaires. We analysed sex-specific relationships of LTPA and OPA (in quartiles) with (1) various cardiovascular risk factors and (2) eating habits using analysis of variance and logistic regression, respectively. RESULTS: In both genders, with and without adjustment for education in addition to age, LTPA was associated negatively with body mass index, body fat, waist circumference, resting heart rate, diastolic blood pressure and triglycerides, and positively associated with high-density lipoprotein cholesterol (all P < or = 0.005). OPA adjusted for age only was positively associated with most cardiovascular risk factors but these associations were not significant after further adjustment on education (except for waist circumference in women). Age- and education-adjusted LTPA was associated with increased frequency of consumption of fruits (odds ratio (OR) = 2.05, 95% confidence interval (CI) 1.68-2.52 in men; OR = 1.90, 95% CI 1.41-2.05 in women) and vegetables (OR = 1.81, 95% CI 1.48-2.21 in men; OR = 2.22, 95% CI 1.66-2.97 in women). CONCLUSIONS: The data emphasise the favourable associations of LTPA, a modifiable behaviour, with various cardiovascular risk factors and healthy eating habits. The results also suggest that the relationships of OPA with cardiovascular risk factors depend, at least in part, on socio-economic status as reflected by educational level.     

Effect of storage time on the retrogradation of banana starch extrudate

Starch was isolated from banana starch and the retrogradation phenomenon was studied using diverse techniques, including an enzymatic measurement. Wide-angle X-ray scattering (WAXS) showed that the sample stored for 7 h presented small peaks and when the storage time increased the peaks increased in intensity. The type of diffraction pattern found in banana extrudates is typical of the A-type crystal polymorph. The crystallinity index from the diffractograms, showed a plateau after approximately 20 h of storage. The short-range order measurement with Fourier transform infrared (FTIR) spectroscopy showed that banana starch retrogradation reached a maximum value at approximately 11 h of storage, a value that agrees with the results obtained with differential scanning calorimetry (DSC), because the maximum enthalpy value (approximately 5 J/g) was calculated in the stored sample for 8 h, without changes in the stored samples for more time. Retrograded resistant starch values did not change after 12 h of storage, obtaining the maximum starch retrogradation level. FTIR, DSC, and the enzymatic technique showed the changes at the molecular level in starch during storage; in the case of WAXS, they determine the long-range order that explains the differences found in the starch retrogradation pattern measurement in banana starch.     

Kepler detected gravity-mode period spacings in a red giant star

Stellar interiors are inaccessible through direct observations. For this reason, helioseismologists made use of the Sun's acoustic oscillation modes to tune models of its structure. The quest to detect modes that probe the solar core has been ongoing for decades. We report the detection of mixed modes penetrating all the way to the core of an evolved star from 320 days of observations with the Kepler satellite. The period spacings of these mixed modes are directly dependent on the density gradient between the core region and the convective envelope.