Comparison of the 5.8S rRNA gene and internal transcribed spacer regions of trichomonadid protozoa recovered from the bovine preputial cavity.

Sequence analysis of the 5.8S rRNA gene and the internal transcribed spacer regions (ITSRs) was used to compare trichomonadid protozoa (n = 39) of varying morphologies isolated from the bovine preputial cavity. A multiple sequence alignment was performed with bovine isolate sequences and other trichomonadid protozoa sequences available in GenBank. As a group, Tritrichomonasfoetus isolates (n = 7) had nearly complete homology. A similarity matrix showed low homology between the T. foetus isolates and other trichomonads recovered from cattle (<70%). Two clusters of trichomonads other than T. foetus were identified. Eighteen isolates comprised 1 group. These isolates shared >99% homology among themselves and with Pentatrichomonas hominis. The other non-T. foetus cluster (n = 14) did not exhibit a high degree of homology (<87%) with other bovine isolates or any of the trichomonad sequences available in GenBank. The sequence homology among isolates in that cluster was >99%, except for 1 isolate that varied from the others in both ITSRs (approximately 2% dissimilarity). Sequence analysis of the 5.8S rRNA gene and ITSRs was useful for comparing trichomonadid protozoa isolated from the bovine preputial cavity and demonstrated that 2 distinct types of trichomonads constituted the non-T. foetus isolates recovered from the bovine preputial cavity.

Cellular mechanisms underlying temperature-induced bleaching in the tropical sea anemone Aiptasia pulchella.

Temperature-induced bleaching in symbiotic cnidarians is a result of the detachment and loss of host cells containing symbiotic algae. We tested the hypothesis that host cell detachment is evoked through a membrane thermotropic event causing an increase in intracellular calcium concentration, [Ca(2+)](i), which could then cause collapse of the cytoskeleton and perturb cell adhesion. Electron paramagnetic resonance measurements of plasma membranes from the tropical sea anemone Aiptasia pulchella and the Hawaiian coral Pocillopora damicornis labeled with 2,2,6,6-tetramethylpiperidine-1-oxyl (TEMPO) revealed no membrane thermotropic event. In addition, intracellular imaging using Fura-2AM as well as labeling anemones with (45)Ca revealed no significant change in [Ca(2+)](i). However, bleaching could be evoked at ambient temperature with 25 mmol l(-1) caffeine without affecting [Ca(2+)](i). [Ca(2+)](i) could be altered with ionomycin in isolated host cells, but ionomycin could not induce bleaching in A. pulchella. As caffeine can affect levels of intracellular protein phosphorylation, the ability of other agents that alter intracellular levels of protein phosphorylation to evoke bleaching was investigated. The protein phosphatase inhibitor vanadate could induce bleaching in A. pulchella. Two-dimensional gels of (32)P-labeled proteins from cold-shocked, caffeine-treated and control anemones show that both temperature shock and caffeine alter the array of phosphorylated host soluble proteins. We conclude that cnidarian bleaching is linked to a temperature-induced alteration in protein phosphorylation.

Identification by subtractive hybridization of sequences specific for Salmonella enterica serovar enteritidis.

Salmonella enterica serovar Enteritidis, a major cause of food poisoning, can be transmitted to humans through intact chicken eggs when the contents have not been thoroughly cooked. Infection in chickens is asymptomatic; therefore, simple, sensitive, and specific detection methods are crucial for efforts to limit human exposure. Suppression subtractive hybridization was used to isolate DNA restriction fragments present in Salmonella serovar Enteritidis but absent in other bacteria found in poultry environments. Oligonucleotide primers to candidate regions were used in polymerase chain reactions to test 73 non-Enteritidis S. enterica isolates comprising 34 different serovars, including Dublin and Pullorum, two very close relatives of Enteritidis. A primer pair to one Salmonella difference fragment (termed Sdf I) clearly distinguished serovar Enteritidis from all other serovars tested, while two other primer pairs only identified a few non-Enteritidis strains. These primer pairs were also useful for the detection of a diverse collection of clinical and environmental Salmonella serovar Enteritidis isolates. In addition, five bacterial genera commonly found with Salmonella serovar Enteritidis were not detected. By treating total DNA with an exonuclease that degrades sheared chromosomal DNA but not intact circular plasmid DNA, it was shown that Sdf I is located on the chromosome. The Sdf I primers were used to screen a Salmonella serovar Enteritidis genomic library and a unique 4,060-bp region was defined. These results provide a basis for developing a rapid, sensitive, and highly specific detection system for Salmonella serovar Enteritidis and provide sequence information that may be relevant to the unique characteristics of this serovar.

Regulation of PGE(2) and PGI(2) release from human umbilical vein endothelial cells by actin cytoskeleton.

Disruption of microfilaments in human umbilical vein endothelial cells (HUVEC) with cytochalasin D (cytD) or latrunculin A (latA) resulted in a 3.3- to 5.7-fold increase in total synthesis of prostaglandin E(2) (PGE(2)) and a 3.4- to 6.5-fold increase in prostacyclin (PGI(2)) compared with control cells. Disruption of the microtubule network with nocodazole or colchicine increased synthesis of PGE(2) 1.7- to 1.9-fold and PGI(2) 1.9- to 2.0-fold compared with control cells. Interestingly, however, increased release of PGE(2) and PGI(2) from HUVEC into the media occurred only when microfilaments were disrupted. CytD treatment resulted in 6.7-fold more PGE(2) and 3.8-fold more PGI(2) released from HUVEC compared with control cells; latA treatment resulted in 17.7-fold more PGE(2) and 11.2-fold more PGI(2) released compared with control cells. Both increased synthesis and release of prostaglandins in response to all drug treatments were completely inhibited by NS-398, a specific inhibitor of cyclooxygenase-2 (COX-2). Disruption of either microfilaments using cytD or latA or of microtubules using nocodazole or colchicine resulted in a significant increase in COX-2 protein levels, suggesting that the increased synthesis of prostaglandins in response to drug treatments may result from increased activity of COX-2. These results, together with studies demonstrating a vasoprotective role for prostaglandins, suggest that the cytoskeleton plays an important role in maintenance of endothelial barrier function by regulating prostaglandin synthesis and release from HUVEC.

Salpingitis in Pekin ducks associated with concurrent infection with Tetratrichomonas sp. and Escherichia coli.

Increased mortality (1.5% per week) and low egg production (5-10% lower than normal) were observed in a flock of domestic breeding Pekin ducks (Anas platyrhynchos). At necropsy, salpingitis and peritonitis were the most significant findings. Histologically, there was accumulation of necrotic debris in the lumen of the oviduct. Numerous bacteria and trichomonads were observed histologically in the lumen of the vagina and occasionally in the shell gland. Escherichia coli and a trichomonad were isolated from the oviduct. The trichomonads were oval (6-8 microm long, 4.5-6 microm wide) and had 4 anterior flagella and an undulating membrane extending over the entire length of the body, finishing in a long posterior flagellum. Morphology was consistent with trichomonads of the genus Tetratrichomonas. Comparative sequence analysis of the 5.8S ribosomal RNA gene and the flanking internal transcribed space regions of the trichomonad isolate did not closely match with available sequences of the same region of other trichomonadid protozoa.

Effects of supplemental instruction on mean test scores and failure rates in medical school courses.

PURPOSE: To determine whether supplemental instruction offered to first-year medical students reduces the number of examination failures. METHOD: A student-run, optional, supplemental-instruction program called the Medical Scholars Program (MSP) was offered at no cost to all first-year students at the University of Southern California School of Medicine in 1994-95. Supplemental instruction was offered in a small-group format in biochemistry, gross anatomy, microanatomy, and physiology. Weekly two-hour sessions were conducted by second-year medical students during the first trimester of the year-1 curriculum. Mean test scores and failure rates for students considered academically at risk and those not at risk were compared between the class entering in 1994 and the classes matriculating during the preceding three years. At-risk students were defined as those with a total Medical College Admission Test score below 26 and a science grade-point average below 3.0. Comparisons were performed using two-tailed t-tests and chi-square tests. RESULTS: Statistically significant increases in mean test scores were achieved on most examinations by the class exposed to the MSP. Failure rates for at-risk students decreased by 46% during the year the MSP was offered. CONCLUSION: Supplemental instruction can significantly improve student performance and therefore retention, particularly among at-risk students.

Neurophysiological Correlates of the Behavioral Response to Light in the Sea Anemone Anthopleura elegantissima.

Neurophysiological responses to light in Anthopleura elegantissima do not involve the ectodermal slow system 1 (SS1). Activities in both the endodermal slow system 2 (SS2) and the through conducting nerve net (TCNN) change when the lighting changes, but the response is not consistent. Thus, photoreception in A. elegantissima probably occurs in the endoderm because SS2 and the TCNN are involved and SS1 is not. We hypothesize either that the photoreception occurs in sensory cells in a local nerve net, with the information then being transmitted to the muscles, or that the muscles themselves are light sensitive. In either case, the TCNN and SS2 become involved after the transduction, and as a consequence--rather than the cause--of muscular activation. The conducting systems of zooxanthellate specimens have a higher frequency of activity than those of apozooxanthellate individuals.