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PubMed2006

Ventricular septal defect and aortic valve regurgitation: pathophysiology and indications for surgery.

As the velocity of a fluid increases a low-pressure zone is created, this is the Venturi effect and it explains the pathogenesis of aortic valve prolapse (AVP) and aortic insufficiency (AI) that is observed in a subset of patients with a ventricular septal defect (VSD). The VSDs complicated by AI are restrictive with high velocity shunting through the VSD, creating a low-pressure zone that impacts the adjacent aortic valve cusp resulting in AVP and subsequent AI. AVP and AI are therefore acquired lesions. AI is absent at birth because the forces necessary to create the low-pressure zone within the restrictive VSD do not exist in utero. The risk of development of AI increases during childhood, peaking at 5 to 10 years of age. VSD closure eliminates the low-pressure zone that is the cause of ongoing aortic valve cusp deformity and, if performed early, prevents development of AI. Patients with a subarterial VSD and AVP should undergo surgery to prevent the development of AI because this complicates about half of subarterial VSDs with AVP and spontaneous closure is rare. Patients with perimembranous VSDs with AVP should be followed with serial echocardiography and undergo VSD closure if more than trivial AI develops.

Aortic Valve Insufficiency↗

PubMed2006

Sperm DNA damage is related to field fertility of semen from young Norwegian Red bulls.

Flow cytometry was utilised for the first time to independently measure five sperm parameters of individual spermatozoa of bull ejaculates to differentiate between outcome successes after artificial insemination (AI). These parameters included plasma membrane and acrosome integrity, mitochondrial functionality and DNA damage measured by sperm chromatin structure assay (SCSA) and terminal deoxynucleotide transferase-mediated dUTP nick end labelling (TUNEL) assays. For each parameter, results of 142 ejaculates (30 bulls) were ranked into three groups according to their flow cytometric measures: (1) ejaculates with the 25% lowest measures; (2) the 50% middle measures; and (3) the 25% highest measures. In total, 20 272 first-service inseminations (18 ;10(6) spermatozoa per AI dose) were performed, where fertility was defined as non-return within 60 days after first insemination. While plasma membrane and acrosome integrity, and mitochondrial functionality were not significantly related to fertility, data from SCSA and TUNEL assays were significantly associated with fertility. Ejaculates in SCSA group 1 had higher odds of AI success (1.07, 95% CI = 1.02-1.12), whereas those in group 3 had lower odds of AI success (0.94, 95% CI = 0.89-0.99), compared with the average odds of all three groups. Ejaculates in group 2 did not have significantly higher odds of AI success compared with the average odds. For TUNEL-positive spermatozoa, the odds of AI success was higher in group 1 compared with the average odds (1.10, 95% CI = 1.02-1.13), whereas odds of AI success in groups 2 and 3 were not significant compared with the average odds. In conclusion, despite the high number of spermatozoa per AI dose from high-quality bulls, both SCSA and TUNEL assays were valuable measures in this study for evaluating sperm quality in relation to fertility after AI.

Animals↗

PubMed1974

Renal hypertension in rats immunized against angiotensin I and angiotensin II.

Rats, actively immunized against angiotensin I (AI) and angiotensin II (AII), were subjected to unilateral renal artery constriction to determine whether the resulting hypertension, which may still ensue in the animal immunized against AII, could be prevented by such combined immunity. Sustained immunity to both AI and AII neither changed preoperative blood pressures of the rats from those of control mock-immunized rats nor altered the incidence or severity of renal dip hypertension. Vascular hyperresponsiveness to small quantities of free angiotensin could not be invoked to explain the hypertension, for there was no significant difference between mock-immunized hypertensive animals, and those remaining normotensive, regarding pressor sensitivity to intravenous AI, AII, renin, and norepinephrine. (AI + AII)-immunized hypertensive rats required AI doses averaging 260 times greater than nonimmune hypertensives to elicit equipressor responses, and were refractory to renin, but not to norepinephrine. Thus, while previous studies have not excluded direct participation of endogenous AI in renal clip hypertension in rats, evidence from our experiments makes it extremely difficult to sustain any pressor function therein for circulating AI or AII. Our results also preclude involvement of AII produced from circulating AI by conversion within arteriolar walls, close to receptor sites, since AI immunity would block this mechanism of action.

Angiotensin II↗

PubMed1997

Papanicolaou smear sensitivity for adenocarcinoma in situ of the cervix. A study of 34 cases.

The sensitivity of cervical smears for adenocarcinoma in situ (AIS) is not known, nor is it known whether false-negative smears are due to sampling or to screening or interpretive errors. In 16 of 34 patients with AIS, 38 negative smears were reported 2 weeks to 7 years before biopsy. Thirty-one of these negative smears were rescreened, and 17 (55%) were retrospectively diagnosed as abnormal. Ten of the 17 had numerous well-preserved AIS cells: 5 with very small, crowded AIS cells, possibly originally mistaken for endometrial cells, and 5 with large groups in which AIS cells resembled reactive endocervical cells. Four smears were confirmed sampling errors. The sensitivity of cervical smears for AIS was 55% to 72%. Improved sampling of the endocervical canal offers cytologists the opportunity to diagnose AIS. This study demonstrates that this opportunity may not be fully exploited. Small "endometrioid" AIS cells and AIS cells resembling reactive endocervical cells may be mistaken for benign cells, thus decreasing sensitivity.

Adenocarcinoma↗

PubMed1999

Expression of the insecticidal bean alpha-amylase inhibitor transgene has minimal detrimental effect on the nutritional value of peas fed to rats at 30% of the diet.

The effect of expression of bean alpha-amylase inhibitor (alpha-AI) transgene on the nutritional value of peas has been evaluated by pair-feeding rats diets containing transgenic or parent peas at 300 and 650 g/kg, respectively, and at 150 g protein/kg diet, supplemented with essential amino acids to target requirements. The results were also compared with the effects of diets containing lactalbumin with or without 0.9 or 2.0 mg bean alpha-AI, levels equivalent to those in transgenic pea diets. When 300 and 650 g peas/kg diet were fed, the daily intake of alpha-AI was 11.5 or 26.3 mg alpha-AI, respectively. At the 300 g/kg level, the nutritional value of the transgenic and parent line peas was not significantly different. The weight gain and tissue weights of rats fed either of the two pea diets were not significantly different from each other or from those of rats given the lactalbumin diet even when this was supplemented with 0.9 g alpha-AI/kg. The digestibilities of protein and dry matter of the pea diets were slightly but significantly lower than those of the lactalbumin diet, probably due to the presence of naturally occurring antinutrients in peas. The nutritional value of diets containing peas at the higher (650 g) inclusion level was less than that of the lactalbumin diet. However, the differences between transgenic and parent pea lines were small, possibly because neither the purified recombinant alpha-AI nor that in transgenic peas inhibited starch digestion in the rat small intestine in vivo to the same extent as did bean alpha-AI. This was the case even though both forms of alpha-AI equally inhibited alpha-amylase in vitro. Thus, this short-term study indicated that transgenic peas expressing bean alpha-AI gene could be used in rat diets at 300 g/kg level without major harmful effects on their growth, metabolism and health, raising the possibility that transgenic peas may also be used at this level in the diet of farm animals.

Animals↗

PubMed1988

Identifying sleep apnea from self-reports.

An apnea score (AS) was developed as a potential screening tool for sleep apnea. This was based on self-report questionnaire responses of 76 sleep disorder center patients and 20 sleep survey volunteers. Twenty volunteers and 23 patients (group I) comprised the initial AS development group. Their questionnaire responses were compared to polysomnographic apnea indexes (AI) and apnea plus hypopnea indexes (AHI). Stepwise multivariate discriminant analysis was used to test whether or not selected group I questionnaire responses could be used to correctly classify respondents into apnea (AI or AHI greater than 5) or nonapnea (AI, AHI less than or equal to 5) groups. Self-reports of "stops breathing during sleep," "loud snoring," and history of adenoidectomy best discriminated normal (AI less than or equal to 5) from apnea (AI greater than 5) cases. The AS derived from group I responses to these three variables was then computed for group II (n = 53). After examination of the AS results, the AS was modified to include just "stops breathing" and "loud snoring" and the AI criterion was raised to 10 per hour. This revised AS correctly identified 100% of the cases with moderate-severe sleep apnea (AI or AHI greater than 40) and 70-76% of all sleep apnea cases with AI or AHI greater than 5. Predictive accuracy was 88% for AI greater than 10. The two questions that comprise the AS should be incorporated into risk appraisal instruments or interviews to screen for sleep apnea.

Adenoidectomy↗

PubMed1997

Using self-reported questionnaire data to prioritize OSA patients for polysomnography.

Many laboratories have large numbers of patients with suspected obstructive sleep apnea (OSA) waiting to be tested. We assessed the use of simple clinical data to detect those patients with an apnea index <20 (low AI) who could be studied less emergently. Using questionnaires completed by patients prior to evaluation, we collected data on 354 consecutive patients (281 males, 73 females; mean age 48.6 years) referred for OSA and assessed with polysomnography (PSG). The questionnaires included the Epworth sleepiness scale (ESS), height, weight, age, and a history of observed apnea. Analysis of receiver operating characteristics curves revealed that both body mass index (BMI) [area under curve = 0.7258, standard error (SE) = 0.03, p < 0.01] and ESS (area under curve = 0.5581, SE = 0.03, p = 0.03) were significantly better than chance alone in detecting people with AI < 20. ESS < or =12 was found in 37.9% of the subjects but 39.6% of those expected to have a low AI using ESS had an AI > or =20. A BMI < or =28 was found in 24.9% of the subjects; 14.8% of those expected to have a low AI using BMI had an AI > or =20. Combining these variables improved accuracy but resulted in smaller groups; a cut-off of ESS < or =12 and BMI < or =28 resulted in a group of 33 (9.3% of subjects), only two (6%) of whom were falsely called low AI. Adding to this the fact that apnea had not been observed resulted in a group of nine patients (2.5% of subjects), none of whom had an AI > or =20. Thus there is a tradeoff; the more variables used, the greater the accuracy but the smaller the percent of cases selected to have low AI. However, in laboratories with hundreds of patients waiting to be tested, any procedure better than chance to help prioritize patients seems worthwhile.

Appointments and Schedules↗

PubMed1997

Effects of equine chorionic gonadotropin, human chorionic gonadotropin, and laparoscopic artificial insemination on embryo, endocrine, and luteal characteristics in the domestic cat.

The effects of gonadotropin treatment and laparoscopic artificial insemination (AI) on embryo quality, serum progesterone and estradiol concentrations, and luteal progesterone content were examined in the domestic cat. These data were compared to similar historical data reported for naturally estrual, mated queens. All queens in this study (n = 32) were treated with eCG followed by 1) natural breeding (eCG-NB), 2) NB and hCG (eCG-NB-hCG), 3) NB and a sham AI procedure (eCG-NB-sham AI), or 4) hCG and actual AI (eCG-hCG-AI). Queens ovulating in response to treatment were ovariohysterectomized, and oviducts and uteri were flushed to collect embryos. Ovarian structures were recorded, corpora lutea (CL) were excised and evaluated for progesterone content, and serum was analyzed for estradiol-17beta and progesterone. Follicle and CL numbers ranged from 0 to 28 and 2 to 42 per cat, respectively, and treatment means did not differ (p > or = 0.05) among groups. Embryos were recovered from oviducts and uterine horns in all treatment groups, and recovery ranged from 60-96%. Mean embryo number per queen ranged from 8.2 +/- 2.6 to 23.2 +/- 3.8 and did not differ (p > or = 0.05) among groups. However, the proportions of unfertilized oocytes were greater (p < 0.05) for groups treated with hCG and/or artificially inseminated, and the proportion of blastocysts produced (31 of 107, 29.0%) was lower (p < 0.05) in the eCG-hCG-AI group than for any other treatment (range, 59 of 116 [50.9%] to 67 of 116 [57.8%]). Not all queens in each group produced good-quality embryos (eCG-NB, 5 of 5; eCG-NB-hCG, 5 of 8; eCG-NB-sham AI, 2 of 5; and eCG-hCG-AI, 3 of 6). Serum progesterone and estradiol-17beta, and total luteal progesterone per ovary did not differ (p > or = 0.05) among treatments. Compared to historical controls (naturally estrual, mated queens), eCG-NB queens produced > 4 times as many good-quality embryos and blastocysts. Similarly, eCG-hCG-AI-treated queens produced > 4 times the number of oocytes and embryos, although a high proportion of these were poor quality and did not develop to blastocysts. Together, these results indicate that queens treated with eCG are capable of consistently producing many good-quality embryos, at least half of which develop to blastocysts in culture. These data support the use of eCG in felids and suggest that other factors are responsible for reduced pregnancy success and small litter sizes following assisted reproduction.

Animals↗

PubMed1998

Apoptotic bodies: a consistent morphologic feature of endocervical adenocarcinoma in situ.

To evaluate the occurrence of apoptotic bodies in endocervical adenocarcinoma in situ (AIS) and investigate the relationship of apoptosis to mitotic activity, we performed counts of apoptotic bodies and mitotic figures in 43 patients with AIS and in a comparable control group with nonneoplastic endocervical glandular epithelium. The ages of the patients with AIS ranged from 27 to 74 years (mean = 40). Mitotic figures were present in all AIS cases, and apoptotic bodies were seen in all but two extremely small lesions. In 28 AIS cases in which lesions were large enough to count 10 consecutive high-power fields (HPF), counts of apoptotic bodies ranged from 1 to 36/10 HPF (mean and median = 16), and counts of mitotic figures ranged from 1 to 53 mitotic figures/10 HPF (mean and median = 18). Counts of apoptotic bodies correlated directly with counts of mitotic figures. The ages of the 28 control patients ranged from 32 to 56 years (mean = 43). Counts of apoptotic bodies in the control cases ranged from 0 to 10 per case (mean = 1.1). Apoptotic bodies were present in only 13 (46%) control cases. The highest counts in these cases ranged from 1 to 6 apoptotic bodies/10 HPF (mean = 2.3). Mitotic figures were present in only 4 (14%) control cases. The counts in these cases ranged from 1 to 3 mitotic figures/10 HPF (mean = 1.8). Counts per 10 HPF were significantly (p < 0.001) more for AIS cases than for controls for apoptotic bodies and mitotic figures. Our results indicate that apoptotic bodies, as well as mitotic figures, occur almost universally in AIS. Both occur significantly more often and in greater numbers in AIS than in nonneoplastic endocervical glandular epithelium. Apoptotic bodies are a consistent morphologic feature of AIS, and their identification may be diagnostically useful.

Adenocarcinoma↗

PubMed1997

Solid viscus injury predicts major hollow viscus injury in blunt abdominal trauma.

BACKGROUND: As nonoperative management of blunt abdominal trauma has become more popular, reliable models for predicting the likelihood of concomitant hollow viscus injury in the hemodynamically stable patient with a solid viscus injury are increasingly important. METHODS: The Pennsylvania Trauma Systems Foundation registry was reviewed for the period from January 1992 to December 1995 for all adult (age > 12 years) patients with blunt trauma and an Abbreviated Injury Scale (AIS) score > or = 2 for a solid viscus (kidney, liver, pancreas, spleen). Patients with an initial systolic blood pressure < 90 mm Hg were excluded. Hollow viscus injuries included only lacerations or perforations of the gallbladder, gastrointestinal tract, or urinary tract. RESULTS: In the 4-year period, 3,089 patients sustained solid viscus injuries, 296 of whom had a hollow viscus injury (9.6%). The mean age was 35.6 years, mean Injury Severity Score was 22.2, and mean Revised Trauma Score was 7.3; 63.3% of the patients were male. A solitary solid viscus injury occurred in 2,437 patients (79%), 177 of whom (7.3%) had a hollow viscus injury. The frequency of hollow viscus injury increased with the number of solid organs injured: 15.4% of patients with two solid viscus injuries (n = 547) and 34.4% of patients with three solid viscus injuries (n = 96) suffered a concomitant hollow viscus injury (p < 0.001 vs. one organ). A hollow viscus injury was 2.3 times more likely for two solid viscus injuries and 6.7 times more likely for three solid viscus injuries compared with a solitary solid viscus injury. For solitary solid viscus injury, the frequency of hollow viscus injury varied little with increasing AIS score (AIS score 2, 6.6%; AIS score 3, 8.2%; AIS score 4, 9.2%; AIS score 5, 6.2%) (p = 0.27 between groups), suggesting that the incidence of hollow viscus injury is related more to the number of solid visceral injuries than the severity of individual organ injury. Also, when the sum of the AIS scores for solid viscus injuries was <6, the mean rate of hollow viscus injury was 7.8%. This increased to 22.8% when the sum of the AIS scores for solid viscus injury was > or =6 (p < 0.001). A pancreatic injury in combination with any other solid viscus injury had a rate of hollow viscus injury of >33%. CONCLUSION: A model of organ injury scaling predicted hollow viscus injury. Multiple solid viscus injuries, particularly pancreatic, or abdominal solid viscus injuries with an AIS score > or = 6, were predictive of hollow viscus injury. Identification of these injury patterns should prompt consideration for early operative intervention.

Abbreviated Injury Scale↗

PubMed2005

Effects of backpack loading on the pulmonary capacities of normal schoolgirls and those with adolescent idiopathic scoliosis.

STUDY DESIGN: A prospective evaluation of the effects of backpack carriage on the pulmonary function of schoolgirls without spinal deformity versus those with adolescent idiopathic scoliosis (AIS). OBJECTIVE: To establish if recommended backpack load limits for normal schoolchildren are also appropriate for study participants with AIS. SUMMARY OF BACKGROUND DATA: The weight of schoolchildren's backpacks are of concern because of effects including compromise of pulmonary function. Impaired pulmonary function is also found with AIS, but the effect of backpack carriage on the respiratory parameters of schoolchildren with AIS has not previously been examined. METHODS: Forced vital capacity (FVC), forced expiratory volume (FEV1), peak expiratory flow (PEF), and forced expiratory flow (FEF25-75%) were recorded in 17 girls (mean age, 12 years) with moderate AIS (Cobb angle, 26 degrees-50 degrees) and 18 girls (mean age, 11 years) without musculoskeletal deformity during carriage of a backpack loaded at 0%, 5%, 7.5%, 10%, 12.5%, and 15% body weight in random order. Absolute values and proportions of reference values were analyzed by repeated-measures analysis of variance. RESULTS.: No interaction between load and group was found, indicating that backpack loading has a similar effect on the pulmonary function of both normal and AIS groups. However, all recorded pulmonary parameters were found to be significantly lower in the AIS than normal group, significantly so for the referenced FVC and PEF. A significant decrease in FVC and FEV1 was found with increasing backpack load, and the load at which these changes were found to be significant was lower than those established in previous studies. CONCLUSIONS: Pulmonary function may be more sensitive to backpack load than previously considered, especially when study participants with AIS are being considered, and the recommended loading limit of 10% body weight may not be applicable to schoolgirls with AIS.

Back↗

PubMed2006

The search for idiopathic scoliosis genes.

STUDY DESIGN: A cohort of 145 patients with adolescent idiopathic scoliosis (AIS) were identified and contacted to determine whether they had a family history of scoliosis. These results were submitted to an internal genealogical database to screen for potential connections to other AIS families. The severity and incidence of AIS in extended family groups were also analyzed. OBJECTIVES: Our objectives were to quantify the genetic effect in AIS, determine the expressivity and penetrance of AIS in large family groupings, and examine larger scoliosis pedigrees for evidence of multiple genes. SUMMARY OF BACKGROUND DATA: Previous reports have suggested an 80% connectedness among scoliosis families, but no clear evidence of multiple genes. It is not known if there are major gene(s). METHODS: A cohort of 145 AIS probands were identified and contacted to ascertain whether they had a family history of AIS. Their medical records and spine radiographs were reviewed to confirm the diagnosis and determine the disease severity. Using an internal genealogical database, the cases were screened for potential connections that would produce larger extended pedigrees. RESULTS: Overall, 131 of the probands were in the database and 127 showed connections to other scoliosis families, a 97% connectedness. These results suggest a major scoliosis gene, as more than 50% of the probands were connected by founders that all resided in England in the mid 1500s. The differences in penetrance (41% vs. 34%) and expressivity (38% vs. 61%) between seemingly unrelated large family groupings might suggest that two different genes are a major influence for AIS in these families. CONCLUSIONS: Nearly all (97%) AIS patients have familial origins. There appears to be at least one major gene, and the differences in penetrance and expressivity in two large unconnected pedigrees might suggest the presence of more than one gene.

Cohort Studies↗

PubMed1994

Accidents and incidents involving patients in a mental health service.

Healthcare givers, and their managers, have a duty to monitor the frequency and types of accidents and untoward incidents which occur to patients in their care. The purpose of this study was to systematically examine accidents and incidents (AIs) involving patients in a mental health service unit. Data were collected on all inpatient and day patient AIs over a 6-month period (January to June 1991). The data were then analysed to identify: (a) the nature of AIs occurring in the unit; (b) the frequencies of the AIs, and (c) the times at which AIs occurred. In the wards for elderly patients, an increase of AIs was noted from 05.00 hours; after this time the frequency of AIs remained fairly constant until 20.00 hours when a decrease occurred. In the other wards, a rise in AIs could be seen from 07.00 hours and a fall occurred after 01.00 hours. However, within the time period of 07.00-01.00 hours, further patterns were noted in the data for all AIs in the wards for non-elderly patients, as well as for incidents involving aggression and self-harm. Possible reasons for these findings are suggested, and recommendations made for the examination of some nursing practices within the unit.

Accidental Falls↗

PubMed1994

Hepatitis C virus chronic infection as a common cause of mixed cryoglobulinaemia and autoimmune liver disease.

OBJECTIVES: Mixed cryoglobulinaemia (MC) and autoimmune chronic hepatitis (AI-CH) are frequently associated with hepatitis C virus (HCV) chronic infection. Because HCV represents a possible common aetiological factor, the aim of the present study is to investigate the clinico-serological alterations of both MC and AI-CH and to verify a possible overlap between these disorders. SETTING: Patients from three tertiary referral centres. SUBJECTS: Two Italian series of 88 MC patients and 30 AI-CH type 1 were studied. RESULTS: MC and AI-CH share several clinico-serological features. The patients' mean age (MC vs. AI-CH: 60 +/- 9 vs. 57 +/- 13 years), disease duration (10.5 +/- 5 vs. 9.6 +/- 6 years), and female/male ratio (3.4 vs. 3.3) were very similar in the two series. Typical hallmarks of MC, i.e. purpura, arthralgias, and weakness, circulating mixed cryoglobulins with rheumatoid factor activity, and hypocomplementemia were also recorded in a significant number of AI-CH patients. Similarly, chronic active hepatitis was found in 68% of MC patients and its histological and serological alterations were comparable with those of AI-CH; moreover, amongst various autoantibodies, antinuclear antibodies and/or anti-smooth-muscle antibodies were detected in half of the cases of MC. Anti-HCV antibodies, detected by second-generation Chiron ELISA and RIBA, were present in a high percentage of both MC and AI-CH (94 vs. 80%), and frequently associated with HCV viraemia (86 vs. 77%). Finally, anti-GOR, the HCV-related autoantibodies, were found in half cases of MC and AI-CH. CONCLUSIONS: On the whole, these data suggest that HCV in combination with other infectious and environmental and genetic factors can trigger a complex immunological disorder with different clinical patterns.

Aged↗

PubMed2002

Synchronisation of oestrus and reproductive performance of dairy cows following administration of oestradiol benzoate or gonadotrophin releasing hormone during a synchronised pro-oestrus.

OBJECTIVE: To compare the use of gonadotrophin releasing hormone (GnRH) and oestradiol benzoate (ODB) administered following a synchronised pro-oestrus on reproductive performance of lactating dairy cows and the submission rates of non-pregnant cows following resynchronisation. DESIGN: Cohort study. PROCEDURE: Lactating Holstein cows enrolled in a controlled breeding program were first treated with an intravaginal progesterone releasing insert (IVP4) for 8 days, 2.0 mg of ODB intramuscular (i.m.) at device insertion (Day 0), an analogue of PGF2alpha at device removal and either 1.0 mg of ODB i.m., 24 h after device removal (ODB group, n = 242), or 0.25 mg of a GnRH agonist (GnRH group, n = 152) injected i.m. approximately 34 h after device removal. Every cow was artificially inseminated between 49 and 56 h after removal of its insert (Day 10). Cows detected in oestrus 1 day after artificial insemination (AI) that were not detected in oestrus on the previous day were re-inseminated on that day. All cows treated on Day 0 were resynchronised for reinsemination by insertion of a used IVP4 device on Day 23. Oestradiol benzoate at a dose of 1.0 mg was administered i.m. at the time of device insertion. Inserts were removed 8 days later (Day 31) and 1.0 mg of ODB was injected i.m. 24 h later. Those cows detected in oestrus between Days 31 and 35 were artificially inseminated. On Day 46 these cows were resynchronised for a third round of AI by insertion of an IVP4 device, used previously to synchronise cows for the first and second rounds of AI, and administration of 1.0 mg of ODB i.m.. Eight days later inserts were removed. Cows detected in oestrus between Days 54 and 58 were artificially inseminated. Bulls were run with the herd between rounds of AI and removed after 21 weeks of mating. RESULTS: Treatment with ODB or GnRH at the first synchronised pro-oestrus did not significantly alter the reproductive performance over three rounds of AI or over a 21-week breeding period. Treatment also did not alter submission rates at the second round of AI or the proportion of non-pregnant and non-return cows ('phantom' cows) detected and did not result in significant differences in concentrations of progesterone in plasma 10 and 18 days after removal of inserts at the first round of AI. Treatment with GnRH reduced the proportion of cows detected in oestrus at the first round of AI (36.2 vs 97.5%; P < 0.001). CONCLUSION: Administration of GnRH compared to ODB at a synchronised pro-oestrus results in similar reproductive performance. Treatment with GnRH reduced the proportion of cows detected in oestrus following treatment. This may offer advantages to the way AI is managed by enabling insemination at a fixed-time and removing the need for the detection of oestrus.

Administration, Intravaginal↗

PubMed2005

Characterization of a thermoacidophilic L-arabinose isomerase from Alicyclobacillus acidocaldarius: role of Lys-269 in pH optimum.

The araA gene encoding L-arabinose isomerase (AI) from the thermoacidophilic bacterium Alicyclobacillus acidocaldarius was cloned, sequenced, and expressed in Escherichia coli. Analysis of the sequence revealed that the open reading frame of the araA gene consists of 1,491 bp that encodes a protein of 497 amino acid residues with a calculated molecular mass of 56,043 Da. Comparison of the deduced amino acid sequence of A. acidocaldarius AI (AAAI) with other AIs demonstrated that AAAI has 97% and 66% identities (99% and 83% similarities) to Geobacillus stearothermophilus AI (GSAI) and Bacillus halodurans AI (BHAI), respectively. The recombinant AAAI was purified to homogeneity by heat treatment, ion-exchange chromatography, and gel filtration. The purified enzyme showed maximal activity at pH 6.0 to 6.5 and 65 degrees C under the assay conditions used, and it required divalent cations such as Mn2+, Co2+, and Mg2+ for its activity. The isoelectric point (pI) of the enzyme was about 5.0 (calculated pI of 5.5). The apparent Km values of the recombinant AAAI for L-arabinose and D-galactose were 48.0 mM (Vmax, 35.5 U/mg) and 129 mM (Vmax, 7.5 U/mg), respectively, at pH 6 and 65 degrees C. Interestingly, although the biochemical properties of AAAI are quite similar to those of GSAI and BHAI, the three AIs from A. acidocaldarius (pH 6), G. stearothermophilus (pH 7), and B. halodurans (pH 8) exhibited different pH activity profiles. Based on alignment of the amino acid sequences of these homologous AIs, we propose that the Lys-269 residue of AAAI may be responsible for the ability of the enzyme to act at low pH. To verify the role of Lys-269, we prepared the mutants AAAI-K269E and BHAI-E268K by site-directed mutagenesis and compared their kinetic parameters with those of wild-type AIs at various pHs. The pH optima of both AAAI-K269E and BHAI-E268K were rendered by 1.0 units (pH 6 to 7 and 8 to 7, respectively) compared to the wild-type enzymes. In addition, the catalytic efficiency (kcat/Km) of each mutant at different pHs was significantly affected by an increase or decrease in Vmax. From these results, we propose that the position corresponding to the Lys-269 residue of AAAI could play an important role in the determination of the pH optima of homologous AIs.

Aldose-Ketose Isomerases↗

PubMed1995

AinS and a new family of autoinducer synthesis proteins.

In Vibrio fischeri, the autoinducer N-3-oxohexanoyl-L-homoserine lactone (AI-1) governs the cell density-dependent induction of the luminescence operon via the LuxR transcriptional activator. The synthesis of AI-1 from bacterial metabolic intermediates is dependent on luxI. Recently, we found a second V. fischeri autoinducer molecule, N-octanoyl-L-homoserine lactone (AI-2), that in E. coli also activates the luminescence operon via LuxR. A locus independent of luxI was identified as being required for AI-2 synthesis. This 2.7-kb ain (autoinducer) locus was characterized by transposon insertion mutagenesis, deletion and complementation analysis, and DNA sequencing. A single 1,185-bp gene, ainS, was found to be the sole exogenous gene necessary for the synthesis of AI-2 in Escherichia coli. In addition, a V. fischeri ainS mutant produced AI-1 but not AI-2, confirming that in its native species ainS is specific for the synthesis of AI-2. ainS is predicted to encode a 45,580-Da protein which exhibits no similarity to LuxI or to any of the LuxI homologs responsible for the synthesis of N-acyl-L-homoserine lactones in a variety of other bacteria. The existence of two different and unrelated autoinducer synthesis genes suggests the occurrence of convergent evolution in the synthesis of homoserine lactone signaling molecules. The C-terminal half of AinS shows homology to a putative protein in Vibrio harveyi, LuxM, which is required for the synthesis of a V. harveyi bioluminescence autoinducer. Together, AinS and LuxM define a new family of autoinducer synthesis proteins. Furthermore, the predicted product of another gene, ainR, encoded immediately downstream of ainS, shows homology to LuxN, which is similarly encoded downstream of luxM in V. harveyi and proposed to have sensor/regulator functions in the bioluminescence response to the V. harveyi auto inducer. This similarity presents the possibility that AI-2, besides interacting with LuxR, also interacts with AinR under presently unknown conditions.

4-Butyrolactone↗

PubMed2002

pfs-dependent regulation of autoinducer 2 production in Salmonella enterica serovar Typhimurium.

Bacterial intercellular communication provides a mechanism for signal-dependent regulation of gene expression to promote coordinated population behavior. Salmonella enterica serovar Typhimurium produces a non-homoserine lactone autoinducer in exponential phase as detected by a Vibrio harveyi reporter assay for autoinducer 2 (AI-2) (M. G. Surette and B. L. Bassler, Proc. Natl. Acad. Sci. USA 95:7046-7050, 1998). The luxS gene product mediates the production of AI-2 (M. G. Surette, M. B. Miller, and B. L. Bassler, Proc. Natl. Acad. Sci. USA 96:1639-1644, 1999). Environmental cues such as rapid growth, the presence of preferred carbon sources, low pH, and/or high osmolarity were found to influence the production of AI-2 (M. G. Surette and B. L. Bassler, Mol. Microbiol. 31:585-595, 1999). In addition to LuxS, the pfs gene product (Pfs) is required for AI-2 production, as well as S-adenosylhomocysteine (SAH) (S. Schauder, K. Shokat, M. G. Surette, and B. L. Bassler, Mol. Microbiol. 41:463-476, 2001). In bacterial cells, Pfs exhibits both 5'-methylthioadenosine (MTA) and SAH nucleosidase functions. Pfs is involved in methionine metabolism, regulating intracellular MTA and SAH levels (elevated levels of MTA and SAH are potent inhibitors of polyamine synthetases and S-adenosylmethionine dependent methyltransferase reactions, respectively). To further investigate regulation of AI-2 production in Salmonella, we constructed pfs and luxS promoter fusions to a luxCDABE reporter in a low-copy-number vector, allowing an examination of transcription of the genes in the pathway for signal synthesis. Here we report that luxS expression is constitutive but that the transcription of pfs is tightly correlated to AI-2 production in Salmonella serovar Typhimurium 14028. Neither luxS nor pfs expression appears to be regulated by AI-2. These results suggest that AI-2 production is regulated at the level of LuxS substrate availability and not at the level of luxS expression. Our results indicate that AI-2-dependent signaling is a reflection of metabolic state of the cell and not cell density.

Bacterial Proteins↗