Short term outcome after left atrial appendage occlusion with the AMPLATZER Amulet and WATCHMAN device: results from the ORIGINAL registry (saxOnian RegIstry analyzinG and followINg left atrial Appendage cLosure).

BACKGROUND: Various randomized multicenter studies have shown that percutaneous left atrial appendage closure (LAAC) is not inferior in stroke prevention compared to vitamin K antagonists (VKA) and can be performed safely and effectively. AIMS: The prospective multicenter ORIGINAL registry in the Free State of Saxony (saxOnian RegIstry analyzinG and followINg left atrial Appendage cLosure) investigated the efficiency and safety of LAAC with Watchman or Amulet device in a real word setting. A special focus was put on the influence of LAAC frequency on periprocedural efficiency and safety. METHODS AND RESULTS: The total of 482 consecutive patients (Abbott Amulet N = 93 and Boston Scientific Watchman N = 389) were included in the periinterventional analyses. After 6 weeks, 353 patients completed the first follow-up including transoesophageal echocardiography (TEE) (73.2%). Successful LAAC could be performed in more than 94%. The complication rate does not significantly differ between device types (p = 0.92) according to Fischer test and comprised 2.2% in the Amulet and 2.3% in the Watchman group. The kind of device and the frequency of LAAC per study center had no influence on the success and complication rates. Device related thrombus could be revealed more frequently in the Watchman group (4.5%) than in the Amulet group (1.4%) but this difference is still not significant in Fisher test (p = 0.14). Same conclusion can be made about residual leakage 1.1% versus 0% [not significant in Fisher test (p = 0.26)]. Dual antiplatelet therapy followed the intervention in 64% and 22% of patients were discharged under a combination of an anticoagulant (VKA/DOAC/Heparin) and one antiplatelet agent. CONCLUSIONS: The ORIGINAL registry supports the thesis from large, randomized trials that LAAC can be performed with a very high procedural success rate in the everyday clinical routine irrespective of the used LAA device (Watchman or Amulet). The postprocedural antithrombotic strategy differs widely among the participating centers. Trial registration Name of the registry: "saxOnian RegIstry analyzinG and followINg left atrial Appendage cLosure", Trial registration number: DRKS00023803; Date of registration: 15/12/2020 'Retrospectively registered'; URL of trial registry record: https://www.drks.de/drks_web/navigate.do?navigationId=trial.HTML&TRIAL_ID=DRKS00023803 .

Long-term symptom improvement and patient satisfaction after AV-node ablation vs. pulmonary vein isolation for symptomatic atrial fibrillation: results from the German Ablation Registry.

BACKGROUND: We aimed to compare patient characteristics and outcome of patients who had either undergone pulmonary vein isolation (PVI) or AV-node ablation (AVN) to control AF-related symptoms. METHODS: From the German Ablation Registry, we analyzed data of 4444 patients (95%) who had undergone PVI and 234 patients (5%) with AVN. RESULTS: AVN patients were on average 10 years older than PVI patients (71 ± 10 vs. 61 ± 10 years, p < 0.001) with 33% aged > 75 years. AVN patients had significantly more cardiovascular comorbidities (diabetes 21% vs. 8%, renal insufficiency 24% vs. 3%, underlying heart disease 80% vs. 36%, severely reduced left ventricular function 28% vs. 1%, all p < 0.001). Significantly more PVI patients had paroxysmal AF (63% vs. 18%, p < 0.001), and more AVN patients had long-standing persistent AF (44% vs. 7%, p < 0.001). At 1-year follow-up, mortality in the AVN group was much higher (Kaplan-Meier estimates 9.8% vs. 0.5%). 20% of PVI patients had undergone another ablation vs. 3% AVN patients (p < 0.001). Symptomatic improvement was equally achieved in about 80%. Re-hospitalization for cardiovascular reasons occurred significantly more often in PVI vs. AVN patients (31% vs. 18%, p < 0.001). CONCLUSION: In the large German Ablation Registry, AVN ablation was performed much less frequently than PVI for symptomatic treatment of AF and typically in older patients with more comorbidity. Symptomatic improvement was similar in both groups. Hospitalizations for cardiovascular reasons were lower in AVN patients despite older age and more cardiovascular comorbidities. 20% of PVI patients had undergone at least one re-ablation.

Effect of Ioxaglate on the cutaneous microcirculation in patients with coronary artery disease: Randomized, double blind, placebo-controlled study.

Radiographic contrast media (RCM) can initiate microcirculatory disorders. This study was performed to investigate effects of Ioxaglate on the cutaneous microcirculation. The investigation was carried out as prospective randomized double-blind comparison in parallel-group design on two groups of n = 10 patients each who had to undergo a diagnostic coronary angiography.The confirmatory parameter of the study was mean erythrocyte capillary velocity [vRBC in mm/sec]. VRBC in the ipsilateral nail-fold capillaries was recorded continuously for 3 min before and 6 min after injection of RCM or isotonic saline solution in the A. axillaris respectively, and was evaluated off-line.VRBC in nailfold capillaries was found to be decreased by Ioxaglate by 34% 150 seconds after injection, while isotonic NaCl solution immediately induced a slight increase of 14%.

[Dyspnea in left-sided heart disease]. / Dyspnoe bei Linksherzerkrankungen.

Shortness of breath (dyspnea) is a common symptom in left-sided heart disease but clinically, patient symptoms show a high variability. Echocardiography is the mainstay for evaluating whether left-sided heart disease is the cause of dyspnea. If left-sided heart failure is diagnosed, this symptom complex must then be subjected to further etiological evaluation. Hypertensive, ischemic and valvular heart diseases are common, as well as atrial fibrillation. If the patient does not have angina pectoris, testing for ischemic heart disease should be done non-invasively by coronary computed tomography or testing for regional myocardial ischemia. Coronary revascularization is indicated only when a prognostically relevant ischemia of more than 10 % of the left ventricle is diagnosed. Diuretics are important for the relief of dyspnea but do not improve the prognosis of patients. In patients with reduced left ventricular function, combination therapy with angiotensin-converting enzyme (ACE) inhibitors, beta blockers and aldosterone antagonists improve the symptoms and prognosis. For treatment of heart failure with preserved ejection fraction evidence-based measures are still lacking. In this case the recommended therapy consists of optimal treatment of comorbidities, regulation of heart rate and blood pressure and participation in structured exercise programs. Angiotensin receptor blockers and aldosterone antagonists can be given in patients with more severe symptoms even though the available data are very sparse.

[Reflex syncope and syncope secondary to orthostatic hypotension]. / Reflexsynkopen und Synkopen bei orthostatischer Hypotonie.

BACKGROUND: Reflex syncope predominantly occurs in younger patients and is the most common type of syncope. Typical contributors to reflex syncope are orthostatic stress, followed by a delayed and inadequate circulatory response consisting of bradycardia (cardioinhibitory type) and hypotension (vasodepressor type). Comparably, syncope may occur after direct activation of the vagus nerve, after emotional distress or pain, and in specific situations, such as coughing and post-micturition. The latter situations are mediated by indirect vagus nerve activation by usually unknown mediators. Syncope mediated by orthostatic hypotension occurs in elderly patients and is mediated by insufficient sympathoadrenergic vasoconstriction, occurring shortly after the onset of the orthostatic situation. DIAGNOSTICS: A thorough examination of the patient history is the mainstay of diagnostics. Specific testing is only required in uncertain and recurrent cases. In addition to standard diagnostics, tilt table testing can be helpful. A negative tilt test is, however, not definitive. Implanted loop recorders are helpful to diagnose the cardioinhibitory component of reflex syncope and are more sensitive than tilt testing. THERAPY: Treatment of both types of syncope consists of avoiding known situations leading to syncope, early reaction to prodromal syndromes, and physical counterpressure manoeuvers. Drug treatment (e.g. alpha-adrenergic agonists and fludrocortisone) are effective only in patients with orthostatic syncope. In selected patients with reflex syncope of a predominantly cardioinhibitory type, pacemaker implantation may be considered in selected patients.

[Anticoagulation in high thromboembolic risk after catheter ablation for atrial fibrillation: results from the German Ablation Registry]. / Antikoagulationssituation bei hohem thromboembolischem Risiko nach Vorhofflimmerablation: Daten des Ablations-Qualitätsregisters.

BACKGROUND: Catheter ablation (CA) for atrial fibrillation (AF) is an effective therapeutic option for the treatment of symptomatic drug-refractory AF. According to current guidelines, the prevention of stroke and embolism is the most important therapeutic goal in AF and the recommendations for anticoagulation (OAC) after successful CA are based upon the CHA2DS2-VASc-Score 3. The aim of this study was to evaluate the use of OAC in patients with a high risk for thromboembolic events 1 year after CA and to identify predictor variables for discontinuation of OAC. METHODS: Between January 2007 and January 2010 13092 patients were enrolled in the study. A total of 52 German electrophysiological centers agreed to participate in this prospective multicenter registry. 41 centers included patients undergoing CA for AF. Analysis included patients who were discharged with OAC after CA and had a CHA2DS2-VASc-Score ≥ 2. A centralized 1 year follow-up (FU) was conducted via telephone. RESULTS: 1300 patients fulfilled the inclusion criteria. One year after CA 51.8 % of these patients were on OAC. Factors significantly associated with discontinuation of OAC included no AF recurrence in FU (adjusted odds ratio (OR): 2.14, [95 % confidence interval (CI): 1.73-2.66], P < 0.001) and paroxysmal AF (OR: 1.53 [95 % CI: 1.29-1.81], P < 0.001). Factors associated with continuation of OAK were patient age (OR per 10 years: 0.79 [95 % CI: 0.68-0.91], P = 0.002), valvular heart disease (OR: 0.67 [95 % CI: 0.48-0.92], P = 0.013), an implanted pacemaker, defibrillator or a cardiac resynchronization therapy system (OR: 0.55 [95 % CI: 0.41-0.74], P < 0.001) and neurological events in hospital or during FU (OR: 0.40 [95 % CI: 0.18-0.88], P < 0.022). CONCLUSION: Almost half of the patients with an indication for OAC are not adequately anticoagulated one year after CA for AF. Paroxysmal AF or freedom from AF is significantly associated with discontinuation of OAC.

[Percutaneous coronary intervention versus bypass surgery in patients with diabetes and multivessel coronary disease. Coronary revascularization after FREEDOM]. / Perkutane koronare Intervention versus Bypass-Operation bei Patienten mit Diabetes und koronarer Mehrgefäßerkrankung. Koronarrevaskularisation nach FREEDOM.

Is coronary revascularization required in a patient with chronic stable coronary artery disease or can optimized medical therapy (OMT) alone be a sufficient alternative? This question has been controversially discussed for non-diabetics as well as for diabetics since the COURAGE and BARI 2D trials. According to our present knowledge, a patient will benefit from coronary revascularization only when either a non-invasive test method, such as single photon emission computed tomography (SPECT) or positron emission tomography (PET) myocardial scintigraphy, stress echocardiography or stress nuclear magnetic resonance imaging, can detect relevant, objective evidence of ischemia >10% of the left ventricular myocardium or when a pathological fractional flow reserve (FFR) <0.80 can be measured in an invasive procedure for an angiographically detectable coronary stenosis. If similar relevant ischemia can be non-invasively or invasively objectified in a patient with chronic stable multivessel coronary artery disease, the often controversially discussed question arises particularly in diabetics whether a percutaneous coronary intervention (PCI) with implantation of drug-eluting stents or coronary artery bypass surgery should be favored. The FREEDOM study (Future Revascularization Evaluation in Patients with Diabetes Mellitus: Optimal Management of Multivessel Disease), published in November 2012, was the first prospective randomized study to examine this issue in diabetic patients with multivessel coronary artery disease. Despite a higher rate of stroke in the surgical cohort, after an average follow-up time of 3.8 years a significant prognostic advantage in favor of bypass surgery was detected for a combined primary endpoint of all-cause mortality, nonfatal myocardial infarction and nonfatal stroke. Thus, in the new ESC guidelines on diabetes, pre-diabetes and cardiovascular diseases developed with the EASD of the European Society of Cardiology and published in 2013, coronary bypass surgery has a class I, level of evidence A recommendation for patients with diabetes mellitus, chronic stable multivessel coronary disease and a synergy between PCI with taxus and cardiac surgery (SYNTAX) score >22. The decision for or against a PCI/stent implantation or coronary bypass surgery in a diabetic patient with chronic stable multivessel coronary artery disease should therefore be made with the patient only after a detailed informed consent discussion and comprehensive explanation of both treatment options. In controversial cases, particularly with an equivocal SYNTAX score around 22, relevant comorbidities or anticipated method-specific complications, a one-stage ad hoc intervention during the diagnostic coronary angiography should be rejected in favor of a two-stage procedure with prior discussion of both treatment options in the heart team comprising noninvasive cardiologists, interventional cardiologists and cardiac surgeons.

[Prevalence of undetected diabetes mellitus in invasive and interventional cardiology. Silent diabetes in the catheterization laboratory]. / Prävalenz unentdeckter Glukosestoffwechselstörungen in der invasiven und interventionellen Kardiologie. "Silent diabetes" im Herzkatheterlabor.

In most randomized controlled trials on revascularization therapy for patients with ischemic coronary artery disease (CAD), the diabetes prevalence ranges between 15% and 35%. However, the true prevalence of diabetes is probably considerably underestimated in these trials. The European heart survey diabetes and the heart published in 2004 supplied strong evidence that there are many additional cases of undetected prediabetics and diabetics in any cardiology patient cohort. The long-term outcome of newly detected diabetics was found to be comparable to patients with already known diabetes mellitus. With this in mind, the Dresden silent diabetes study investigated the prevalence of undetected diabetes mellitus by oral glucose tolerance testing (OGTT) and comparative HbA1c sampling in 1,015 patients admitted for coronary angiography. Patients with known diabetes were excluded from the study.According to the OGTT only 513 patients (51%) were classified with normal glucose tolerance (NGT), 10 (1%) with isolated impaired fasting glucose (IFG), 349 (34%) with impaired glucose tolerance (IGT) and 143 (14%) were diagnosed with newly detected diabetes mellitus (DM). According to the HbA1c measurements 588 patients (58%) were classified as normal, 385 (38%) as borderline and only 42 (4%) were diagnosed with diabetes (DM). There was a significant correlation between the extent of CAD and glycemic status as defined by the OGTT. The number of patients with IGT and diabetes increased with the extent of CAD (IGT group p<0.001, diabetes group p=0.01). However, no such correlation was observed when glycemic status was defined by HbA1c testing.Based on these results an OGTT should be routinely performed in patients with known or suspected coronary artery disease undergoing coronary angiography for diagnosis of diabetes, as HbA1c measurements alone appear to miss a substantial proportion of patients. These findings are of high clinical relevance with regard to optimal coronary revascularization procedure chosen in catheterization laboratories, preferably drug-eluting stents in cases of diabetes mellitus or newly detected diabetes mellitus and preferably coronary bypass surgery in diabetics with multi-vessel disease and high SYNTAX scores.

[Outpatient rehabilitation after myocardial infarction or for heart failure]. / Ambulante Rehabilitation nach Herzinfarkt und bei Herzinsuffizienz.

Reducing cardiac mortality and improving quality of life are the main objectives of cardiac rehabilitation. In recent years, outpatient rehabilitation within easy patient reach has achieved the same status as inpatient rehabilitation. Outpatient rehabilitation permits close involvement of the patient's family and social environment, thus easing reintegration into everyday life. However, the health care system is not yet utilizing outpatient rehabilitation to its full potential. This contribution illustrates the principles of rehabilitation following myocardial infarction or for heart failure in an outpatient setting, as well as its potential and future development.

Oral glucose tolerance test and HbA1c for diagnosis of diabetes in patients undergoing coronary angiography: [corrected] the Silent Diabetes Study.

AIMS/HYPOTHESIS: The primary aim of this study was to compare the results of HbA(1c) measurements with those of an OGTT for early diagnosis of 'silent diabetes' in patients with coronary artery disease (CAD) undergoing angiography without prediagnosed diabetes. A secondary aim was to investigate the correlation between the extent of CAD and the glycaemic status of the patient. METHODS: Data from 1,015 patients admitted for acute (n = 149) or elective (n = 866) coronary angiography were analysed. Patients with known diabetes were excluded from the study. Using the OGTT results, patients were classified as having normal glucose tolerance (NGT), impaired fasting glucose (IFG), impaired glucose tolerance (IGT) or diabetes. According to the results of the HbA(1c) measurements, patients were classified into three groups: normal (HbA(1c) <5.7% [<39 mmol/mol]), borderline (HbA(1c) 5.7-6.4% [39-47 mmol/mol]) and diabetes (HbA(1c) ≥6.5% [≥48 mmol/mol]). RESULTS: Based on the OGTT, 513 patients (51%) were classified with NGT, 10 (1%) with IFG, 349 (34%) with IGT and 149 (14%) were diagnosed with diabetes. According to HbA(1c) measurements, 588 patients (58%) were classified as normal, 385 (38%) as borderline and 42 (4%) were diagnosed with diabetes. The proportion of patients with IGT and diabetes increased with the extent of CAD (IGT ρ = 0.14, p < 0.001, diabetes ρ = 0.09, p = 0.01). No differences in HbA(1c) were seen among the groups with different extents of CAD (p = 0.652). CONCLUSIONS/INTERPRETATION: An OGTT should be performed routinely for diagnosis of diabetes in patients with CAD undergoing coronary angiography, since HbA(1c) measurement alone appears to miss a substantial proportion of patients with silent diabetes. A limitation of the study is that the OGTT was not performed before the angiography.

Influence of radiographic contrast media on myocardial tissue oxygen tension: NaCl-controlled, randomised, comparative study of iohexol versus iopromide in an animal model.

It was tested whether a bolus injection of 10 ml radiographic contrast medium (iopromide vs. iohexol), compared to a 10 ml NaCl bolus and administered into the left anterior descending artery (LAD) of farm pigs, influenced the tissue pO2 in the territory of this artery. The radiographic contrast media and the NaCl bolus were given in randomised order. The mean pO2 LAD fell from initially 40.3+/-10.9 mmHg to a minimal value of 22.5+/-8.9 mmHg 241+/-44 sec after injecting the iopromide bolus, with this result representing a mean decrease of 44.2% (p=0.0003). The initial pO2 (baseline) was reached again after approximately 10 minutes. The mean pO2 LAD fell from the initial value of 34.5+/-14.6 mmHg to a minimal value of 29.4+/-13.9 mmHg 171.7+/-11.9 sec after injection of the iohexol bolus, with this result representing a mean decrease of 14.8% (p=0.0003). The baseline pO2 was reached again after approx. 5 minutes. The drop in the pO2 after iopromide administration was significantly larger than that after iohexol (p=0.0001), and also the time after which the baseline pO2 is reached again was considerably shorter for iohexol (p=0.001). The two radiographic contrast media did not influence the tissue pO2 in either the territory of the right coronary artery or in skeletal muscle. Injection of a NaCl bolus into the LAD influenced neither the tissue pO2 of the territory of the LAD nor that of the RCA or of the skeletal muscle. The tissue temperature, heart rate and the systolic and diastolic blood pressure were not affected during the three injections. Injection of radiographic contrast media into a coronary artery can lead to a distinct, local microcirculatory impairment in the myocardial territory supplied by this artery. In this case, the extent of the microcirculation impairment seems to depend not only on the viscosity of the contrast media but rather also on its chemotoxicity.

Influence of two non-ionic radiographic contrast media with different osmolalities on coagulation in invasive cardiology. A prospective, randomised comparative study.

PURPOSE: To investigate the influence of two non-ionic radiographic contrast media with different osmolality on thrombocytic function and the plasmatic coagulation system. MATERIAL AND METHODS: The study was carried out as a randomised, prospective, comparative study with two contrast media in a heart catheter laboratory. RESULTS: Activating influences on platelet aggregation, procoagulatory or profibrinolytic functions or injury to the endothelium could be ruled out. Apparently, also differences in substance properties, such as the media's ionic character or osmolality had no demonstrable influence on the interaction with haemostatis and blood vessels. An adjuvant, antithrombotic therapy was carried out, which consisted of platelet aggregation inhibitors and heparins. CONCLUSION: Our findings agree with the results of recent clinical trials, which demonstrated no relevant disadvantage of non-ionic contrast media as regards thrombotic complications.

Effect of an ionic compared to a non-ionic X-ray contrast agent on platelets and coagulation during diagnostic cardiac catheterisation.

The aim of the present study was to evaluate the effects of ionic (ioxaglate) and non-ionic (iopromide) contrast media on haemostatic parameters ex vivo. In 40 patients undergoing coronary angiography, platelet function (platelet reactivity and serotonin concentration) and coagulation markers [thrombin-antithrombin III complexes, prothrombin fragments (F1+2) and the D-dimers] were measured. The use of an ionic X-ray contrast agent (XCA) (ioxaglate) in diagnostic cardiac catheterisation angiography is associated with lower thrombin generation and lower activation of the platelet system than when a non-ionic XCA is employed (iopromide). The results thus confirm the results of various in vitro studies and animal investigations.

Treatment of atrial fibrillation in open heart surgery--the potential role of microwave energy.

Treatment of atrial fibrillation by creating linear lesions in patients undergoing heart surgery (mitral valve replacement or coronary bypass grafting) receives increasing interest. Conventional rhythm surgery require tremendous operative effort and increases the risks of perioperative complications. For that reason various alternative methods for creating linear lesion are under investigation. This paper discusses several techniques for creation of transmural lesions in open heart surgery, e.g. ultra sound ablation, high current DC shocks, laser ablation, cryoablation, radio frequency ablation, cooled radio frequency ablation, and microwave ablation.

Intraoperative microwave ablation for curative treatment of atrial fibrillation in open heart surgery--the MICRO-STAF and MICRO-PASS pilot trial. MICROwave Application in Surgical treatment of Atrial Fibrillation. MICROwave Application for the Treatment of Atrial Fibrillation in Bypass-Surgery.

Concomitant microwave ablation for curative treatment of atrial fibrillation (AF) was performed in 18 patients with history of chronic atrial fibrillation and indication for open heart surgery, 11 patients with mitral valve replacement and 7 patients with coronary artery bypass grafting. There were no perioperative complications. During the postoperative period most of the patients had intermittent AF, they received low dose Sotalol therapy and electric cardioversions. Up to now seven patients have reached follow-up day 90. One patient has persistent AF. Two patients had typical atrial flutter that was electrically converted to sinus rhythm (SR), isthmus ablation is planned. The other four patients have SR, one patient without cardioversions. These four patients show recovered atrial function with observed A-wave for transmitral flow. Under visual guidance the continuous atrial lesion lines could be induced effectively and safely by the intraoperative device Lynx.

Pacing and ICD therapy in Germany (West and East) before and after reunification in 1989.

In 1989 the two German countries, FRG and GDR, were reunited after 50 years of political separation. During this time, these countries underwent independent, and in parts quite different, developments. While the reunification has had less effect on the overall situation in the Western part of the country, the Eastern part has experienced considerable changes, including the health service. In the field of pacing and ICD therapy, this study finds that 8 years after the reunification, the German federal states of both parts of the country have converged in the field of pacing and ICD therapy.

A prothrombin gene mutation is significantly associated with venous thrombosis.

This case-control study examined the prevalence of a prothrombin gene mutation in the 3'-untranslated region (UTR) first reported by Poort et al in Dutch subjects with a history of venous thrombosis and in matched control subjects without a history of thrombosis. We tested the hypothesis that the presence of the 3'UTR prothrombin mutation would convey a higher risk of venous or arterial thrombosis and therefore would be found in a higher-than-normal percentage of subjects with a history of thrombosis. Our study included 100 subjects: 50 with a history of thrombosis (21 with venous thrombosis and 29 with arterial thrombosis, who had been recruited from an anticoagulation clinic) and 50 control subjects without a history of thrombosis. DNA from these subjects was analyzed by polymerase chain reaction and agarose gel electrophoresis. We found a statistically significant increase in the prevalence of the 3'UTR mutation in subjects with a history of venous thrombosis compared with subjects without thrombosis. The prevalence of the 3'UTR prothrombin mutation was 19% (4/21;3 heterozygous and 1 homozygous) in subjects with a history of venous thrombosis, 0% (0/29) in subjects with a history of arterial thrombosis, and 2% (1/50) in control subjects (P < .0245, by Fisher's exact test for comparison of subjects with versus those without a history of venous thrombosis). The G-->A mutation at nucleotide 20,210 in the 3'UTR was confirmed by direct DNA sequencing. The similar increased prevalence of the 3'UTR mutation in subjects with venous thrombosis in our population and in the Dutch population studied by Poort et al suggests that this mutation is an important risk factor for venous thrombosis in the general white population.

Isolation of Cryptococcus neoformans chromosome-specific probes using expressed sequence tags.

Clinical and environmental isolates of Cryptococcus neoformans exhibit a high degree of karyotypic variability. Analysis of the molecular basis of karyotypic differences requires a large set of chromosome-specific probes. We have determined the chromosomal distribution of a set of randomly selected C. neoformans cDNA clones and have explored the feasibility of identifying these clones through partial DNA sequencing. Forty-four randomly selected cDNA clones were labelled and hybridized to PFGE blots of C. neoformans. Expressed sequence tags were generated by sequencing the 5'-end of each clone. Thirty-five clones hybridized to single bands on PFGE blots. At least seven chromosomes were recognized by these probes. Homology searches identified potential homologs of several groups of proteins not previously studied in C. neoformans. PFGE hybridization and sequencing of random cDNA clones is an efficient method for identifying chromosomal-specific probes in fungi that lack extensive sets of genetic markers.

Genetic relatedness of Cryptococcus neoformans clinical isolates grouped with the repetitive DNA probe CNRE-1.

Cryptococcus neoformans isolates from eight patients with cryptococcal infection were previously assigned into three groups on the basis of repetitive DNA probe (CNRE-1) restriction fragment length polymorphisms. These groups accounted for a disproportionate number of recent clinical isolates in New York City. To further examine the genetic relatedness of isolates within and across CNRE-1 groups, the DNA sequence of the 779-base URA5 gene from each strain was amplified and sequenced. The number of nucleotide differences occurred in the third codon position or in introns. Pairwise comparisons revealed average nucleotide differences within a CNRE-1 group of 4.8 +/- 2.6 (n = 8) and between CNRE-1 groups of 21.9 +/- 7.0 (n =20) (P <0.001) Analysis of URA5 sequences defined three groups that were congruent with those defined by CNRE-1 restriction fragment length polymorphisms. PCR amplification of an rDNA intergenic spacer revealed conservation of the intergenic spacer length within groups. Electrophoretic karyotyping did not distinguish between two isolates in each of two CNRE-1 groups. DNA from all isolates studied hybridized to an alpha mating type-specific probe. We interpret these results as suggesting a clonal population structure for some pathogenic isolates of C. neoformans in New York City.