Circulation dynamics of Salmonella enterica subsp. enterica ser. Gallinarum biovar Gallinarum in a poultry farm infested by Dermanyssus gallinae.

Dermanyssus gallinae (Mesostigmata: Dermanyssidae, De Geer, 1778) is an ectoparasite of poultry, suspected to play a role as a vector of Salmonella enterica subsp. enterica ser. Gallinarum. Despite an association between them being reported, the actual dynamics in field remain unclear. Therefore, the present study aimed to confirm the interactions among mites, pathogen and chickens. The study was carried out in an industrial poultry farm infested by D. gallinae, during an outbreak of fowl typhoid. The presence of S. Gallinarum in mites was assessed and quantified by a semi-nested polymerase chain reaction (PCR) and real-time PCR, respectively, in mites collected during two subsequent productive cycles and the sanitary break. The anti-group D Salmonella antibodies were quantified by an enzyme-linked immunosorbent assay. During the outbreak and the sanitary break, S. Gallinarum was constantly present in mites. In the second cycle, scattered positivity was observed, although hens did not exhibit signs of fowl typhoid, as a result of the vaccination with BIO-VAC SGP695 (Fatro, Ozzano Emilia Bo, Italy). The data strongly suggest that D. gallinae acts as reservoir of S. Gallinarum, thus allowing the pathogen to persist in farms. Furthermore, the present study has highlighted the interactions among D. gallinae, S. Gallinarum and hens with respect to enhancing the mite-mediated circulation of S. Gallinarum in an infested poultry farm.

The septins: roles in cytokinesis and other processes.

The septins are a novel family of proteins that were first recognized in yeast as proteins associated with the neck filaments. Recent work has shown that septins are also present in other fungi, insects, and vertebrates. Despite the apparent differences in modes of cytokinesis amongst species, septins appear to be essential for this process in both fungal and animal cells. The septins also appear to be involved in various other aspects of the organization of the cell surface.

Effects of smoking and different BMI cutoff points on surgical site infection after elective open ventral hernia repair.

PURPOSE: BMI > 40 kg/m2 and smoking have been identified as risk factors for surgical site infections (SSIs) after ventral hernia repair, however, the relationship with lower BMI values has not been described. The objective of this study was to analyze the relationship between different BMI thresholds, smoking, and SSI after open ventral hernia repair. METHODS: All patients who underwent an elective non-emergent open ventral hernia repair with mesh were extracted from the 2011 to 2016 NSQIP database. Bivariate, multivariate logistic regression, and Classification and Regression Tree (CART) analyses were used. RESULTS: Of 55,240 patients, 2,620 (4.7%) developed SSIs (superficial: 58.5%, deep:27%, organ-space: 16%). BMI (OR: 1.035; 95% CI:1.03-1.04; p < 0.001) and smoking (OR:1.51; 95% CI:1.37-1.67; p < 0.001) were identified in logistic regression analysis as the two most modifiable risk factors independently associated with SSIs. CART analysis demonstrated that the lowest SSI rate belonged to non-smokers with BMI < 24.2 kg/m2 (1.9%), and the highest SSI to smokers with BMI > 42.3 kg/m2 (12%). Between these values, there was a stepwise increase in SSI rate as BMI increased, while smoking added additional risk in each group. CONCLUSION: Following open hernia repair, the association between SSI and being overweight starts at a BMI of 24.2, a threshold lower than previously described. The risk of SSI increases in a stepwise fashion as BMI increases and is augmented by smoking. Future studies are needed to determine if SSI reduction can be achieved with a combination of smoking cessation and weight loss using these BMI thresholds.

Outcomes of concomitant mesh placement and intestinal procedures during open ventral hernia repair.

PURPOSE: The concept of mesh use during open ventral hernia repair with a concomitant intestinal procedure remains controversial and it remains unclear whether the SSI profile of contaminated fields in this setting more closely resembles clean-contaminated or dirty wounds. METHODS: Patients who underwent an open ventral hernia repair and intestinal procedures were extracted from the ACS-NSQIP database. Data analysis was performed for mesh versus no mesh groups in aggregate and matched cohorts. The 30-day outcomes including mortality, morbidity, surgical site infections (SSI), readmission, reoperation, and length of stay were compared between the groups. RESULTS: Of 5104 patients in the mesh group, 3297 patients were matched 1:1 with patients without mesh. Mesh placement was associated with higher overall morbidity (35.1% vs. 29.8%; p < 0.001), overall SSI (27.1% vs. 18%; p < 0.001), deep SSI (5.9% vs. 4.3%; p = 0.003), organ-space SSI (6.8% vs. 5.5%; p = 0.027), reoperation (9.9% vs. 8.2%; p = 0.016), readmission (16.7% vs. 12.7%; p < 0.001), and longer hospital stays (10.9 ± 15.2 vs. 9.7 ± 10.7; p < 0.001). When mesh was used, the SSI profile of contaminated fields was similar to dirty wounds (26.4% vs. 27.5%; p = 0.702), rather than clean-contaminated fields (26.4% vs. 19.2%; p < 0.001). CONCLUSION: Concomitant bowel procedure and mesh placement at the time of open ventral hernia repair was associated with worse outcomes. The SSI profile of the contaminated fields was more similar to dirty wounds. When deciding whether to place mesh during ventral hernia repairs with concomitant bowel procedures, strong consideration should be given to the increased risk of short-term complications versus the potential long-term benefits.

A septin-based hierarchy of proteins required for localized deposition of chitin in the Saccharomyces cerevisiae cell wall.

Just before bud emergence, a Saccharomyces cerevisiae cell forms a ring of chitin in its cell wall; this ring remains at the base of the bud as the bud grows and ultimately forms part of the bud scar marking the division site on the mother cell. The chitin ring seems to be formed largely or entirely by chitin synthase III, one of the three known chitin synthases in S. cerevisiae. The chitin ring does not form normally in temperature-sensitive mutants defective in any of four septins, a family of proteins that are constituents of the "neck filaments" that lie immediately subjacent to the plasma membrane in the mother-bud neck. In addition, a synthetic-lethal interaction was found between cdc12-5, a temperature-sensitive septin mutation, and a mutant allele of CHS4, which encodes an activator of chitin synthase III. Two-hybrid analysis revealed no direct interaction between the septins and Chs4p but identified a novel gene, BNI4, whose product interacts both with Chs4p and Cdc10p and with one of the septins, Cdc10p; this analysis also revealed an interaction between Chs4p and Chs3p, the catalytic subunit of chitin synthase III. Bni4p has no known homologues; it contains a predicted coiled-coil domain, but no other recognizable motifs. Deletion of BNI4 is not lethal, but causes delocalization of chitin deposition and aberrant cellular morphology. Overexpression of Bni4p also causes delocalization of chitin deposition and produces a cellular morphology similar to that of septin mutants. Immunolocalization experiments show that Bni4p localizes to a ring at the mother-bud neck that lies predominantly on the mother-cell side (corresponding to the predominant site of chitin deposition). This localization depends on the septins but not on Chs4p or Chs3p. A GFP-Chs4p fusion protein also localizes to a ring at the mother-bud neck on the mother-cell side. This localization is dependent on the septins, Bni4p, and Chs3p. Chs3p, whose normal localization is similar to that of Chs4p, does not localize properly in bni4, chs4, or septin mutant strains or in strains that accumulate excess Bni4p. In contrast, localization of the septins is essentially normal in bni4, chs4, and chs3 mutant strains and in strains that accumulate excess Bni4p. Taken together, these results suggest that the normal localization of chitin synthase III activity is achieved by assembly of a complex in which Chs3p is linked to the septins via Chs4p and Bni4p.

Cell growth control: little eukaryotes make big contributions.

The story of rapamycin is a pharmaceutical fairytale. Discovered as an antifungal activity in a soil sample collected on Easter Island, this macrocyclic lactone and its derivatives are now billion dollar drugs, used in, and being evaluated for, a number of clinical applications. Taking advantage of its antifungal property, the molecular Target Of Rapamycin, TOR, was first described in the budding yeast Saccharomyces cerevisiae. TORs encode large, Ser/Thr protein kinases that reside in two distinct, structurally and functionally conserved, multi-protein complexes. In yeast, these complexes coordinate many different aspects of cell growth. TOR complex 1, TORC1, promotes protein synthesis and other anabolic processes, while inhibiting macroautophagy and other catabolic and stress-response processes. TORC2 primarily regulates cell polarity, although additional readouts of this complex are beginning to be characterized. TORC1 appears to be activated by nutrient cues and inhibited by stresses and rapamycin; however, detailed mechanisms are not known. In contrast, TORC2 is insensitive to rapamycin and physiological regulators of this complex have yet to be defined. Given the unsurpassed resources available to yeast researchers, this simple eukaryote continues to contribute to our understanding of eukaryotic cell growth in general and TOR function in particular.

Bud8p and Bud9p, proteins that may mark the sites for bipolar budding in yeast.

The bipolar budding pattern of a/alpha Saccharomyces cerevisiae cells appears to depend on persistent spatial markers in the cell cortex at the two poles of the cell. Previous analysis of mutants with specific defects in bipolar budding identified BUD8 and BUD9 as potentially encoding components of the markers at the poles distal and proximal to the birth scar, respectively. Further genetic analysis reported here supports this hypothesis. Mutants deleted for BUD8 or BUD9 grow normally but bud exclusively from the proximal and distal poles, respectively, and the double-mutant phenotype suggests that the bipolar budding pathway has been totally disabled. Moreover, overexpression of these genes can cause either an increased bias for budding at the distal (BUD8) or proximal (BUD9) pole or a randomization of bud position, depending on the level of expression. The structures and localizations of Bud8p and Bud9p are also consistent with their postulated roles as cortical markers. Both proteins appear to be integral membrane proteins of the plasma membrane, and they have very similar overall structures, with long N-terminal domains that are both N- and O-glycosylated followed by a pair of putative transmembrane domains surrounding a short hydrophilic domain that is presumably cytoplasmic. The putative transmembrane and cytoplasmic domains of the two proteins are very similar in sequence. When Bud8p and Bud9p were localized by immunofluorescence and tagging with GFP, each protein was found predominantly in the expected location, with Bud8p at presumptive bud sites, bud tips, and the distal poles of daughter cells and Bud9p at the necks of large-budded cells and the proximal poles of daughter cells. Bud8p localized approximately normally in several mutants in which daughter cells are competent to form their first buds at the distal pole, but it was not detected in a bni1 mutant, in which such distal-pole budding is lost. Surprisingly, Bud8p localization to the presumptive bud site and bud tip also depends on actin but is independent of the septins.

A method to study the rapid phosphorylation-related modulation of neutral trehalase activity by temperature shifts in yeast.

Heat shock enhanced the synthesis of neutral trehalase in growing cells of Saccharomyces cerevisiae, as detected by immunological methods. The activity of the enzyme was measured in extracts obtained by two methods: cells were either harvested by filtration and subsequent disruption with glass beads at 0-4 degrees C or immediately frozen with liquid nitrogen in the presence of Triton X-100, followed by thawing at 30 degrees C. The first procedure yielded artificially high activities of neutral trehalase in heat-shocked cells due to rapid (less than 1 min) activation during handling at 4 degrees C before homogenization. Activity of the enzyme in these homogenates decreased 75-90% upon a treatment with alkaline phosphatase, indicating that activation was due to phosphorylation. The second procedure yielded low trehalase activities for heat-shock treated cells, much higher activities for cells shifted back for some seconds to 27 degrees C, and very low activities again for cells shifted from 27 to 40 degrees C for a second time. Thus, permeabilization of cells following rapid freezing in Triton X-100 is a method of choice to study post-translational modulation of the neutral trehalase of S. cerevisiae by phosphorylation and dephosphorylation.

Acquisition of thermotolerance in Saccharomyces cerevisiae without heat shock protein hsp 104 and in the absence of protein synthesis.

Acquisition of thermotolerance in response to a preconditioning heat treatment at 40 degrees C was studied in mutants of the yeast Saccharomyces cerevisiae lacking a specific heat shock protein or the ability to synthesize proteins at 40 degrees C. A mutant carrying a deletion of heat shock protein hsp 104 and the corresponding wildtype strain were both highly sensitive to heat stress at 50.4 degrees C without preconditioning but both acquired almost the same level of thermotolerance after 60 min of preconditioning. Both strains showed equal induction of trehalose-6-phosphate synthase and accumulated equal levels of trehalose during the treatment. The conditional mutant ts--187 synthesized no proteins during the preconditioning heat treatment but nevertheless acquired thermotolerance, albeit to a lesser degree than the corresponding wildtype strain. Induction of trehalose-6-phosphate synthase and accumulation of trehalose were reduced to a similar extent. These results show that acquisition of thermotolerance and accumulation of trehalose are closely correlated during heat preconditioning and are modulated by protein synthesis but do not require it.

Expression of a functional barley sucrose-fructan 6-fructosyltransferase in the methylotrophic yeast Pichia pastoris.

The cDNA encoding sucrose-fructan 6-fructosyltransferase (6-SFT) from barley (Hordeum vulgare) has been expressed in the methylotrophic yeast Pichia pastoris, using a translational fusion into vector pPICZ alphaC, containing the N-terminal signal sequence of Saccharomyces cerevisiae alpha-factor to allow entry into the secretory pathway. Transformed Pichia produced and secreted a functional 6-SFT which had characteristics similar to the barley enzyme, but had a pronounced additional 1-SST activity when incubated with sucrose.

Heat shock induces enzymes of trehalose metabolism, trehalose accumulation, and thermotolerance in Schizosaccharomyces pombe, even in the presence of cycloheximide.

Exponentially growing cells of the fission yeast, Schizosaccharomyces pombe, contained virtually no trehalose at 27 degrees C but rapidly accumulated large quantities during heat shock at 40 degrees C. Activities of trehalose-6-phosphate synthase and trehalase also increased upon heat shock. Thermotolerance of the cells, measured as survival at 52 degrees C, increased in parallel to trehalose accumulation and decreased in parallel to the trehalose levels when cells were shifted back to 27 degrees C. Trehalose levels, activities of enzymes of trehalose metabolism and thermotolerance strongly increased upon heat shock even in the presence of cycloheximide, indicating that none of these effects requires protein synthesis. The data support the hypothesis that trehalose acts as a thermoprotectant in Schizosaccharomyces pombe.

Saccharomyces cerevisiae mitochondria can synthesise FMN and FAD from externally added riboflavin and export them to the extramitochondrial phase.

Evidence is given that mitochondria isolated from Saccharomyces cerevisiae can take up externally added riboflavin and synthesise from it both flavin mononucleotide (FMN) and flavin adenine dinucleotide (FAD) probably due to the existence of the mitochondrial riboflavin kinase already reported and the novel mitochondria FAD synthetase. Moreover Saccharomyces cerevisiae mitochondria can export the newly synthesised flavin derivatives to the extramitochondrial phase. This has been proven to take place with 1:1 stoichiometry with riboflavin decrease outside mitochondria, thus showing that flavin traffic occurs across the mitochondrial membranes.

Myocardial contusion in the stable patient: what level of care is appropriate?

To evaluate the significance of myocardial contusion, we evaluated 243 stable patients hospitalized for blunt chest trauma between 1982 and 1986. The groups were identified according to results of radionuclide angiography, mean injury severity score (ISS), and outcome. Group I (n = 71; mean ISS = 12.7) patients were those without myocardial contusion by radionuclide angiography. Two patients with cardiac complications were in this group. The patients with myocardial contusion were divided into two groups. Group II (n = 69; ISS = 19.5) patients had myocardial contusion as an isolated injury, and group III (n = 103; ISS = 30.9) patients had myocardial contusion and injury to at least one other organ system. Three patients from group II had cardiac complications. Eleven patients from group III had cardiac complications. There were no significant differences between the cardiac complication rate in the three groups, and each complication was present when the patient arrived in the emergency department. The predicted mortality rate based on ISS was 10% to 20% for patients with myocardial contusion, whereas the observed mortality rate for the groups (II and III) overall was 0.58%. We conclude that in the stable trauma patient myocardial contusion (1) does not by itself increase the risk of complication, (2) does not necessitate intensive care unit monitoring, (3) should be devalued when computing ISS scores, (4) may account for lengthy and often unnecessary hospitalization, and (5) in patients at risk for complications may be identified by ECG abnormalities on arrival to the emergency department.

Preoperative cardiac risk assessment: an updated approach.

HYPOTHESIS: We provide an updated algorithm for approaching preoperative cardiac risk assessment in patients undergoing noncardiac surgery. DESIGN: A National Library of Medicine PubMed literature search was performed dating back to 1985 using the keywords "preoperative cardiac risk for noncardiac surgery." This search was restricted to English language articles involving human subjects. RESULTS: Patient-specific and operation-specific cardiac risk can be determined clinically. Patients with major cardiac risk factors have a high incidence of perioperative cardiac complications, whereas the risk is less than 3% for low-risk patients. For intermediate-risk patients, no prospective randomized studies demonstrate the efficacy of noninvasive stress testing (dipyridamole thallium or dobutamine echocardiography) or of subsequent coronary revascularization for preventing perioperative cardiac complications. Recent studies demonstrate that perioperative beta-blockade significantly reduces the adverse cardiac event rate in intermediate-risk patients. CONCLUSIONS: Most patients with high cardiac risk should proceed with coronary angiography. Patients with low cardiac risk can proceed to surgery without noninvasive testing. For intermediate-risk patients, consideration may be given to further stress testing prior to surgery; however, in most patients, proceeding to surgery with perioperative beta-blockade is an acceptable alternative.

Gallstone pancreatitis. The role of preoperative endoscopic retrograde cholangiopancreatography.

OBJECTIVES: To evaluate the efficacy of endoscopic retrograde cholangiopancreatography (ERCP) and laparoscopic cholecystectomy in patients with gallstone pancreatitis and to determine criteria predictive of common bile duct stones (CBDS). DESIGN: Retrospective chart review. PATIENTS: Seventy-one consecutive patients with gallstone pancreatitis. MAIN OUTCOME MEASURES: Identification and endoscopic management of CBDS, complications, and mortality. RESULTS: Preoperatively, ERCP revealed CBDS in seven of 22 patients and postoperatively, in five of six patients. All stones were successfully removed. Laboratory values and common bile duct dilatation on admission did not predict CBDS. Persistent hyperamylasemia (> 150 U/L) and persistent hyperbilirubinemia (> 29.07 mumol/L [1.7 mg/dL]) were associated with CBDS on ERCP or intraoperative cholangiography. All five patients with cholangitis underwent ERCP, and CBDS were found and removed in four. There were no deaths and there was a 7% complication rate. CONCLUSIONS: Gallstone pancreatitis can be effectively managed by selective ERCP, endoscopic sphincterotomy, and laparoscopic cholecystectomy. Preoperative ERCP can be restricted to patients with cholangitis, persistent hyperbilirubinemia, or persistent hyperamylasemia.

Simplified admission criterion for predicting severe complications of gallstone pancreatitis.

HYPOTHESIS: Simple admission criteria (white blood cell count, > or =14. 5 x 10(9)/L; blood urea nitrogen level, > or =4.3 mmol/L [> or =12 mg/dL]; heart rate, > or =100 beats per minute; and serum glucose level, > or =8.3 mmol/L [> or =150 mg/dL]) are better predictors of severe complications of gallstone pancreatitis than an Acute Physiology and Chronic Health Evaluation II (APACHE II) score of 5 or greater, a modified Imrie (Glasgow) score of 3 or greater, and a biliary Ranson score of 3 or greater. DESIGN: A prospective consecutive case study. SETTING: A university-affiliated, urban, public hospital. PATIENTS: Ninety-two consecutive patients (77 women and 15 men, aged 18 to 76 years [mean age, 39 years]) with gallstone pancreatitis. Seventy-seven patients were Hispanic. MAIN OUTCOME MEASURES: Major local and systemic complications requiring intensive care unit care, and death. RESULTS: Fourteen patients (15%) had severe complications with a mortality of 2%. On univariate analysis, a white blood cell count of 14.5 x 10(9)/L or more (P =.03), a serum glucose level of 8. 3 mmol/L or more (> or =150 mg/dL) (P<.001), an APACHE II score of 5 or greater (P =.008), a modified Imrie score of 3 or greater (P<.001), and a biliary Ranson score of 3 or greater (P =.03) were statistically associated with the development of severe complications; whereas a blood urea nitrogen level of 4.3 mmol/L or more (> or =12 mg/dL) and a heart rate of 100 beats per minute or more were not. On multivariate analysis, only a serum glucose level of 8. 3 mmol/L or more (> or =150 mg/dL) was predictive of adverse events (P<. 001). CONCLUSIONS: Glucose level (> or =8.3 mmol/L [> or =150 mg/dL]) is the best single admission predictor of severe complications of gallstone pancreatitis and is superior to an APACHE II score of 5 or greater, a modified Imrie score of 3 or greater, and a biliary Ranson score of 3 or greater.

Endovascular vs open abdominal aortic aneurysm repair: a comparison of cardiac morbidity and mortality.

HYPOTHESIS: Adverse cardiac event rates following endovascular abdominal aortic aneurysm (EAAA) and open abdominal aortic aneurysm (OAAA) repair are similar. We also hypothesized that the Eagle criteria (Q wave on electrocardiogram, diabetes, angina, congestive heart failure, age >70 years, and ventricular ectopy) are useful predictors of cardiac events in patients undergoing EAAA repair. DESIGN: Prospective (patients undergoing EAAA repair) and retrospective (patients undergoing OAAA repair). SETTING: Public teaching and Veterans Affairs medical centers. PATIENTS: Eighty-three EAAA and 63 OAAA repairs. MAIN OUTCOME MEASURES: Myocardial infarction, congestive heart failure, and cardiac death. RESULTS: Patients with EAAA were older (73 vs 68 years, P=.003). There were no differences in the mean number of Eagle criteria (1.2 vs 1.3), cardiac event rates (6% vs 4.8%), or mortalities (3.6% vs 4.8%). Within the EAAA group, congestive heart failure (P=.005) and Q wave on electrocardiogram (P=.006) were the only predictors of cardiac events. CONCLUSIONS: Patients undergoing OAAA and EAAA repair had similar cardiac event rates and mortality. In patients undergoing EAAA repair, history of congestive heart failure and Q wave on electrocardiogram were predictors of cardiac events.

Trends in cardiac morbidity and mortality after endoluminal abdominal aortic aneurysm repair; discussion 999-1000.

HYPOTHESIS: The adverse cardiac event rate following endoluminal abdominal aortic aneurysm (EAAA) repair has decreased as experience in performing the procedure has increased. Aneurysm complexity affects the rate of adverse cardiac events. DESIGN AND PATIENTS: Data from 173 consecutive patients undergoing EAAA repair from 2 successive periods were compared. There were 82 patients in the early group (group 1) and 91 patients in the later group (group 2). MAIN OUTCOME MEASURES: Myocardial infarction, congestive heart failure, unstable angina, major dysrhythmias, death. RESULTS: The cardiac event rate was 8.5% for group 1 vs 16.5% for group 2 (P =.16). Predictors of adverse cardiac events on multivariate analysis were the use of 4 or more graft extensions (P =.04), female sex (P =.01), and number of Eagle risk factors (P<.001). There were 2 postoperative deaths (2.4%) in group 1 and 4 (4.4%) in group 2 (P =.7). CONCLUSIONS: Following EAAA repair: (1) adverse cardiac events were found to correlate with use of 4 or more graft extensions, female sex, and the number of Eagle risk factors; (2) cardiac morbidity and mortality remain significant despite greater experience and improved technology; and (3) operative mortality remains acceptably low.

Fibrin glue inhibits intra-abdominal adhesion formation.

To determine the effect of fibrin glue on intra-abdominal adhesion formation, 45 rats were randomized to three groups. Each animal received two adhesion models. Group 1 received no further treatment. Groups 2 and 3 had the adhesion models covered with fibrinogen from fresh frozen plasma (1.77 g/L) and cryoprecipitate (23.6 g/L), respectively. In group 1, 13 of 15 rats had high-grade adhesions in both models. In group 2, high-grade adhesions were noted in nine of 15 rats in model 1 and in 12 of 15 rats in model 2. In group 3, however, high-grade adhesions were seen in only three of 15 rats in model 1, with 11 rats having no adhesions, and in only two of 15 rats in model 2. Histologic analysis suggested accelerated healing in group 3. We conclude that (1) fibrin glue inhibits intra-abdominal adhesion formation in rats, (2) the inhibitory action is dependent on the fibrinogen concentration of the fibrin glue, and (3) adhesions are reduced by fibrin glue regardless of whether the peritoneal defect is closed.

A constitutive, heat shock-activated neutral trehalase occurs in Schizosaccharomyces pombe in addition to the sporulation-specific acid trehalase.

Trehalase was studied in Schizosaccharomyces pombe cells growing vegetatively on minimal medium and in sporulating cultures. Acid trehalase activity, measured at pH 4.2, was absent in vegetative cells and occurred only in asci, indicating that this activity represented the sporulation-specific trehalase reported previously. In contrast, neutral trehalase, measured at pH 6.0, was constitutively present in vegetative cells during the exponential and stationary growth phase as well as in asci. In vegetative cells, neutral trehalase did not sediment with cell walls, suggesting a cytoplasmic localization. Its activity increased ten-fold when growing cells were subjected to heat treatment of 2 h. Neutral trehalase from heat-treated cells had a pH optimum of 6.0 and was almost completely inhibited by 3 mM ZnCl2. Acid trehalase activity could be measured in intact asci, indicating that it is localized in the ascus cell walls, while neutral trehalase was not detectable in intact asci and appeared to be present primarily in the walls of ascospores and in the ascus epiplasm.