Mycobacterium bovis and M. caprae in Bulgaria: insight into transmission and phylogeography gained through whole-genome sequencing.

BACKGROUND: This study aimed to characterize recent Mycobacterium bovis/M. caprae isolates from Bulgaria by whole-genome sequencing (WGS) to gain a first insight into their molecular diversity, transmission, and position within the global phylogeography of this important zoonotic species. RESULTS: The isolates were obtained from cattle in diverse locations of Bulgaria in 2015-2020 and were identified by microbiological and PCR assays. WGS data were used for phylogenetic analysis that also included M. bovis global dataset. Thirty-seven M. bovis/caprae isolates from Bulgaria were studied and 34 of them were SNP genotyped. The isolates were subdivided into 3 major phylogenetic groups. Type Mbovis-13 (Eu2 complex [western Europe and northern Africa]) included one isolate. Mbovis-37 type included 5 isolates outside of known clonal complexes. The Bulgarian M. caprae isolates formed a sub-group within the Mcaprae-27B cluster which also included 22 M. caprae isolates from Poland, Spain, Germany, and the Republic of Congo. The Bulgarian M. caprae isolates share their latest common ancestors with Spanish isolates. The Mbovis-37 group shares a distant common ancestor (pairwise distance 22-29 SNPs) with an isolate from Poland but was very distant (> 200 SNPs) from the rest of the tree. The Mbovis-13 group shares a common ancestor with two human isolates from Germany. Phylogeographically, both M. bovis clades had limited circulation in northeastern Bulgaria while the majority of the studied isolates (M. caprae) were from central and western provinces. A phylogenetic network-based analysis demonstrated that 11 Bulgarian isolates were separated by 1 to 6 SNPs within four clusters, mostly forming pairs of isolates. CONCLUSION: The obtained WGS analysis positioned the Bulgarian isolates within the global phylogeography of M. bovis/M. caprae. Hypothetically, the observed phylogenetic diversity may not have resulted from livestock trade routes, but instead may reflect the deeply rooted M. bovis/M. caprae phylogeography of Europe. A high level of genetic divergence between the majority of the studied isolates suggests limited active transmission of bTB in Bulgaria during the survey period. At the same time, a possibility of the endemic presence of circulating bTB strains in the form of the latent persistent disease cannot be ruled out.

First Insight into Diversity of Minisatellite Loci in Mycobacterium bovis/M. caprae in Bulgaria.

The aim of this study was to assess the diversity of minisatellite VNTR loci in Mycobacterium bovis/M. caprae isolates in Bulgaria and view their position within global M. bovis diversity. Forty-three M. bovis/M. caprae isolates from cattle in different farms in Bulgaria were collected in 2015-2021 and typed in 13 VNTR loci. The M. bovis and M. caprae branches were clearly separated on the VNTR phylogenetic tree. The larger and more geographically dispersed M. caprae group was more diverse than M. bovis group was (HGI 0.67 vs. 0.60). Overall, six clusters were identified (from 2 to 19 isolates) and nine orphans (all loci-based HGI 0.79). Locus QUB3232 was the most discriminatory one (HGI 0.64). MIRU4 and MIRU40 were monomorphic, and MIRU26 was almost monomorphic. Four loci (ETRA, ETRB, Mtub21, and MIRU16) discriminated only between M. bovis and M. caprae. The comparison with published VNTR datasets from 11 countries showed both overall heterogeneity between the settings and predominantly local evolution of the clonal complexes. To conclude, six loci may be recommended for primary genotyping of M. bovis/M. caprae isolates in Bulgaria: ETRC, QUB11b, QUB11a, QUB26, QUB3232, and MIRU10 (HGI 0.77). VNTR typing based on a limited number of loci appears to be useful for primary bTB surveillance.

Detection of HEV RNA Using One-Step Real-Time RT-PCR in Farrow-to-Finish Pig Farms in Bulgaria.

(1) Background: HEV is a zoonotic, foodborne pathogen. It is spread worldwide and represents a public health risk. The aim of this study was to evaluate the presence of HEV RNA in farrow-to-finish pig farms in different regions of Bulgaria; (2) Methods: Isolation of HEV RNA from pooled samples of feces was performed using a QIAamp® Viral RNA Mini Kit followed by HEV RNA detection using a single-step real-time RT-PCR with primers and probes targeting the ORF 3 HEV genome; (3) Results: HEV RNA was detected in 12 out of 32 tested farms in Bulgaria (37.5%). The overall percentage of HEV-positive pooled fecal samples was 10.8% (68 of 630 samples). HEV was detected mostly in pooled fecal samples from finisher pigs (66/320, 20.6%) and sporadically from dry sows (1/62, 1.6%) and gilts (1/248, 0.4%); (4) Conclusions: Our results confirm that HEV circulates in farrow-to-finish pig farms in Bulgaria. In our study, we found HEV RNA in pooled fecal samples from fattening pigs (4-6-months age), shortly before their transport to the slaughterhouse indicating a potential risk to public health. The possible circulation of HEV throughout pork production requires monitoring and containment measures.

Effect of sports training on morphological characteristics in Bulgarian female tennis players.

Abstract.

First insight into phylogeography of Mycobacterium bovis and M. caprae from cattle in Bulgaria.

Bovine tuberculosis (bTB) represents a significant economic burden to the agriculture. In spite of decades of the control program, Mycobacterium bovis infection levels in cattle in Bulgaria continued to rise over recent years. In order to gain a better understanding of the M. bovis diversity, we used spoligotyping for strain differentiation and the data were compared to the international databases Mbovis.org and SITVIT2 for shared type and clade assignment. Study sample included 30 M. tuberculosis complex isolates from cattle originating from different regions of Bulgaria. The isolates were subdivided by spoligotyping into 4 spoligotypes: 2 types shared by 20 and 8 isolates and 2 singletons. SITVIT2-defined types SIT645 and SIT647 belonged to the common and classical bovine ecotype M. bovis (9 isolates) while types SIT120 and SIT339 belonged to the M. caprae ecotype (21 isolates). A certain phylogeographic gradient of the spoligotypes and clades at the within-country level was observed: M. caprae was prevalent in the central/southwestern, while classical M. bovis in the northeastern Bulgaria. Whereas all four types have global or European circulation, neither was described in the neighboring Balkan countries. M. caprae isolates identified in this study mostly belong to the Central/Eastern European cluster. In summary, this study provided a first insight into phylogeography of M. bovis in Bulgaria and described, for the first time, M. caprae as an important infectious agent of bTB in this country.

Correlation of cerebellar volume with eyeblink conditioning in healthy subjects and in patients with cerebellar cortical degeneration.

In the present study, acquisition and timing of conditioned eyeblink responses (CRs) were correlated with magnetic resonance imaging (MRI)-based cerebellar volume both in healthy human subjects and patients with cerebellar disease. Thirty-three healthy subjects and 25 patients with pure cortical cerebellar degeneration participated. Cerebellar volumes were measured for the cortex of the anterior lobe, the cortex of the posterior lobe, the white matter of the cerebellum and the cerebrum based on 3D MR images. CR parameters were assessed in a standard delay paradigm. In healthy subjects CR acquisition was significantly related to the volume of the grey matter of the posterior lobe, but neither to the volume of the grey matter of the anterior lobe, nor to the cerebellar white matter and nor to the cerebral volume. As expected, CR acquisition and volume of the cortex of the posterior lobe showed age-related decline in the controls. Furthermore, CR acquisition was significantly reduced in patients with cerebellar degeneration compared to controls. In the cerebellar group, however, no significant correlations between CR acquisition and any of the cerebellar volumes were observed. Floor effects are most likely responsible for this observation. Although CRs occurred significantly earlier in cerebellar patients compared to controls, no significant correlations between CR timing parameters and any of the cerebellar volumes were observed. Extending previous findings in healthy human subjects, age-related decline of the cerebellar cortex of the posterior lobe was related with a reduction of CR acquisition. Findings provide further evidence that the cerebellar cortex plays an important role in the acquisition of eyeblink conditioning in humans.

Cognitive functions in patients with MR-defined chronic focal cerebellar lesions.

The aim of the present study was to examine cognitive functions in a group of chronic patients with focal cerebellar lesions. Both effects of localization (anterior vs. posterior lobe) and side (left vs. right cerebellar hemisphere) were of interest. Fourteen patients with infarctions within the territory of the posterior inferior cerebellar artery (PICA) and seven patients with infarctions within the territory of the superior cerebellar artery (SCA) participated. The affected lobules and nuclei were assessed based on 3D MR imaging. The right cerebellar hemisphere was affected in eight PICA and two SCA patients, the left hemisphere in six PICA and four SCA patients. One SCA patient revealed a bilateral lesion. In order to study possible lateralization of functions, subjects performed a language task as well as standard neglect and extinction tests. Moreover, two tests of executive functions were applied. There were no significant group differences apart from a verbal fluency task, in which all cerebellar patients - but especially those with right-sided lesions - were impaired. Voxel-based lesion-symptom mapping (VLSM) revealed that a lesion of the right hemispheric lobule Crus II was associated with impaired performance in the verbal fluency task. In sum, the results showed preserved cognitive abilities in chronic cerebellar patients apart from impairments of verbal fluency in patients with right-cerebellar lesions. The latter findings are in line with the assumption that the right posterolateral cerebellar hemisphere supports functions associated with verbal fluency.

Timing of conditioned eyeblink responses is impaired in cerebellar patients.

In the present study, timing of conditioned eyeblink responses (CRs) was investigated in cerebellar patients and age-matched controls using a standard delay paradigm. Findings were compared with previously published data of CR incidences in the same patient population (Gerwig et al., 2003; Timmann et al., 2005). Sixteen patients with pure cortical cerebellar degeneration (spinocerebellar ataxia type 6 and idiopathic cerebellar ataxia), 14 patients with lesions within the territory of the superior cerebellar artery, and 13 patients with infarctions within the territory of the posterior inferior cerebellar artery were included. The affected cerebellar lobules and possible involvement of cerebellar nuclei were determined by three-dimensional magnetic resonance imaging (MRI) in patients with focal lesions (n = 27). Based on a voxel-by-voxel analysis, MRI lesion data were related to eyeblink conditioning data. CR incidence was significantly reduced, and CRs occurred significantly earlier in patients with cortical cerebellar degeneration and lesions of the superior cerebellum compared with controls. Incidence and timing of CRs was not impaired in patients with lesions restricted to the posterior and inferior cerebellum. Voxel-based MRI analysis revealed that cortical areas within the anterior lobe (Larsell lobule HV) were most significantly related to timing deficits, whereas reduced CR incidences were related to more caudal parts (lobule HVI) of the superior cerebellar cortex. The present data suggest that different parts of the superior cerebellar cortex may be involved in the formation of the stimulus association and appropriate timing of conditioned eyeblink responses in humans. Extracerebellar premotoneuronal disinhibition, however, is another possible explanation for changes in CR timing.

Involvement of the human cerebellum in short-term and long-term habituation of the acoustic startle response: a serial PET study.

OBJECTIVE: Numerous studies have shown an involvement of the human cerebellum in motor learning, but little is known about the role of the cerebellum in learning of unspecific aversive reactions. The present study sought to distinguish which areas of the human cerebellum and brain-stem are involved in short-term habituation (STH) and long-term habituation (LTH) of the acoustic startle response. METHODS: On 5 consecutive days 42 acoustic startle stimuli were applied each day in 8 male healthy subjects. On the first and on the fifth day of the experiment [15O]H2O PET scans were performed. RESULTS: Electromyographic recordings revealed a significant decrease of the startle response within each day (STH) and across the 5 days of the experiment (LTH). On both days a decrease of regional cerebral blood flow (rCBF) across PET scans was found in the medial cerebellum most probably reflecting reduced sensory feedback during STH. Between days an increase of rCBF in the dorsomedial pons, in the mesencephalon and in an area of the medial cerebellum was observed. These activations may reflect increased inhibition of the startle response during LTH and correspond to previous animal lesion studies. Furthermore, during LTH an increase of rCBF within the lateral cerebellum in lobule HVI/Crus I was detected. CONCLUSIONS: These results suggest that distinct parts of the medial and lateral cerebellum are involved in habituation of the acoustic startle response. Lobule HVI/Crus I most likely plays a more general role in implicit learning processes considering its involvement in several conditioning paradigms. SIGNIFICANCE: The results of the present study contribute to the understanding of cerebellar involvement in learning of unspecific aversive reactions.

Extinction of conditioned eyeblink responses in patients with cerebellar disorders.

Extinction of conditioned eyeblink responses (CRs) was analyzed in sixteen patients with pure cortical cerebellar degeneration, 14 patients with lesions within the territory of the superior cerebellar artery (SCA), 13 patients with infarctions within the territory of the posterior inferior cerebellar artery (PICA) and 45 age-matched controls. Three-dimensional (3D) magnetic resonance (MRI) data sets were acquired in patients with focal lesions to identify affected cerebellar lobules and possible involvement of nuclei. Eyeblink conditioning was performed using a standard delay protocol. At the end of the experiment 10 CS-alone trials were presented as extinction trials. Controls showed significant effects of extinction that is a significant decline comparing CR-incidences in the extinction trials and the last block of 10 trials of the paired trials. In the group of all cerebellar patients, however, no significant effects of extinction were observed. In patients with unilateral lesions effects of extinction were present on the unaffected, but not on the affected side. Deficits of extinction were observed in PICA and SCA patients both with and without involvement of cerebellar nuclei. Extending previous reports in cerebellar patients the present findings show that the ipsilateral cerebellar hemisphere contributes to extinction of conditioned eyeblink responses in humans. It cannot be ruled out, however, that impaired acquisition affected the extinction results.

Cerebellar involvement in verb generation: an fMRI study.

A possible role of the human cerebellum in the generation of verbs corresponding to presented nouns has been suggested. Previous functional brain imaging studies have compared generation of verbs with the reading of nouns as a measure of verb generation. In the present fMRI study involving healthy human subjects, the effects of speech articulation and motor imagery associated with verb production were investigated in greater detail. Generation of verbs to visually presented nouns was compared to a condition in which subjects read those same verbs that had been individually generated by each subject. Activation in lobule HVI/Crus I of the right cerebellar hemisphere was found as a measure of verb generation. In contrast, reading of verbs as a measure of speech articulation evoked cerebellar activations in both left and right paravermal lobule VI. These results suggest an involvement of the right lateral cerebellar hemisphere in linguistic functions during verb generation. Alternatively, effects of inner speech could also possibly explain the results.

Functional recovery of children and adolescents after cerebellar tumour resection.

This study examined whether lesions to the cerebellum obtained in early childhood are better compensated than lesions in middle childhood or adolescence. Since cerebellar lesions might affect motor as well a cognitive performance, posture, upper limb and working memory function were assessed in 22 patients after resection of a cerebellar tumour (age at surgery 1-17 years, minimum 3 years post-surgery). Working memory was only impaired in those patients who had received chemo- or radiation therapy. Postural sway was enhanced in 64% of the patients during dynamic posturography conditions, which relied heavily on vestibular input for equilibrium control. Upper limb function was generally less impaired, but 54% of the patients revealed prolonged deceleration times in an arm pointing task, which probably does not reflect a genuine cerebellar deficit but rather the patients' adopted strategy to avoid overshooting. Age at surgery, time since surgery or lesion volume were poor predictors of motor or cognitive recovery. Brain imaging analysis revealed that lesions of all eight patients with abnormal posture who did not receive chemo- and/or radiation therapy included the fastigial and interposed nuclei (NF and NI). In patients with normal posture, NI and NF were spared. In 11 out of 12 patients with abnormal deceleration time, the region with the highest overlap included the NI and NF and dorsomedial portions of the dentate nuclei in 10 out of 12 patients. We conclude that cerebellar damage inflicted at a young age is not necessarily better compensated. The lesion site is critical for motor recovery, and lesions affecting the deep cerebellar nuclei are not fully compensated at any developmental age in humans.

Vermal atrophy of alcoholics correlate with serum thiamine levels but not with dentate iron concentrations as estimated by MRI.

Chronic alcohol consumption is frequently accompanied by cerebellar degeneration. The exact aetiology of alcoholic cerebellar degeneration is still a matter of debate. The aim of the present study was to investigate whether patients with chronic alcohol consumption exhibit a decrease in dentate nuclei intensity as measured by MRI, and if so, whether this decrease correlates with cerebellar atrophy as revealed by MR imaging or with clinical signs of cerebellar ataxia. A decrease in dentate nuclei intensity would indirectly indicate that iron accumulation, and therefore, oxidative stress may play a role in alcoholic cerebellar degeneration. MRI of 45 alcoholics and 44 age and sex-matched healthy control subjects was performed using a 3D-T1-weighted fast low angle shot (FLASH) echo sequence. Signal intensities of the dentate nuclei and cerebellar white matter were bilaterally measured. Planimetric measurements of cerebellar size were performed using a 3D-T1-weighted magnetization prepared rapid acquisition gradient echo (MPRAGE) sequence. Results demonstrated that dentate nuclei intensity was not significantly decreased in patients with chronic alcohol consumption (mean +/- SD signal intensity 65.36 +/- 13.0) if compared with control subjects (mean +/- SD signal intensity 68.95 +/- 9.4) (p = 0.15). Dentate nuclei intensity did not correlate with cerebellar size neither in control subjects nor in alcoholics. In contrast, vitamin B1 level correlated with cerebellar size in alcoholics even if the vitamin B1 concentration was within normal values (r = 0.344, p = 0.028). These results support the view that thiamine deficiency rather than direct neurotoxic effects of alcohol is the main causative factor for the development of alcoholic cerebellar degeneration.

Behavioral and affective changes in children and adolescents with chronic cerebellar lesions.

The aim of the present study was to investigate if clinically relevant affective or behavioral changes as described in adults in the cerebellar affective syndrome by Schmahmann and Sherman [The cerebellar cognitive affective syndrome, Brain 121 (1998) 561-579] are likely to occur as a long-term sequelae of cerebellar vermis lesions in children. Site and extent of the vermal lesion were defined on the basis of individual 3D-MRI scans and lesion data were correlated with behavioral and affective changes. Affect and behavior were assessed in children after cerebellar tumor surgery by means of experimenter ratings based on the description of the cerebellar affective syndrome and free ratings by the patients and their parents. Twelve children and adolescents with a former cerebellar astrocytoma surgery without subsequent radiation or chemotherapy participated. Detailed analysis of individual 3D-MR images revealed that lesions affected the vermis in nine children. Experimenter ratings according to Schmahmann revealed no relevant problems in patients. In five out of nine patients with vermal affection somewhat increased thoughtful, anxious or aggressive behavior was reported by patients and parents. In conclusion, minor behavioral and affective changes were present in a subset of children with chronic vermal lesions.

Brain representation of hemifield stimulation in poststroke visual field defects.

BACKGROUND AND PURPOSE: Plasticity in extended, parallel, or reciprocal operating networks is well recognized. Changes in neuronal activity after lesions to distinct localized structures, such as the primary visual cortex, are less well characterized. We investigated the cortical reorganization in patients with poststroke visual field defects using blood oxygen level-dependent functional MRI. METHODS: Brain activation was measured in 7 patients with a single occipital cortical lesion and partially recovered hemianopia and in 7 age-matched control subjects. Differences in activation between rest and visual hemifield stimulation were assessed with statistical parametric mapping (SPM'99). RESULTS: In normal subjects, significant activation was found in the contralateral primary visual cortex and bilaterally in the extrastriate cortex. During hemifield stimulation of the unaffected side of stroke patients, a similar pattern was found compared with that seen in control subjects. During stimulation of the hemianopic side, bilateral activation was seen within the extrastriate cortex, stronger in the ipsilateral hemisphere. The primary visual cortex was not significantly activated in either hemisphere during stimulation of the hemianopic side. CONCLUSIONS: Visual field defects after stroke are associated with bilateral activation of the extrastriate visual cortex. This pattern of activation indicates altered neuronal activity in the visual system. Further investigation is necessary to determine the relationship between functional reorganization and recovery of lost visual function after poststroke hemianopia.

Strong evidence for association between the dystrobrevin binding protein 1 gene (DTNBP1) and schizophrenia in 488 parent-offspring trios from Bulgaria.

BACKGROUND: The gene encoding the dystrobrevin binding protein (DTNBP1) has been implicated in the pathogenesis of schizophrenia by several association studies. We tried to replicate these findings in a sample of 488 parent-proband trios recruited in Bulgaria. Probands had a diagnosis of schizophrenia (n = 441) or schizoaffective disorder (n = 47). METHODS: We genotyped eight single nucleotide polymorphisms within the gene, four of which had been reported in previous studies, and four identified as informative by our group through direct screening of the gene and genotyping in a sample of cases and control subjects. RESULTS: A significant excess of transmissions was observed for two of the markers, p1635 and p1757, (p =.0009 and.0013, respectively). Analysis of two-, three-, and four-marker haplotypes produced numerous positive results, with six (4% of the total combinations) at p <.001. CONCLUSIONS: These results provide strong support for DTNBP1 as a susceptibility gene for schizophrenia; however, different haplotypes seem to be associated in different studies.

Major psychiatric disorders and the serotonin transporter gene (SLC6A4): family-based association studies.

The serotonin transporter (5-HTT) is a suitable candidate gene to test for involvement in the pathogenesis of major psychiatric disorders. We used the method of family-based controls to test for association between disease and a variable number tandem repeat (VNTR) in intron 2 of the gene, which has received support for involvement in the pathogenesis of several psychiatric disorders. We analysed 413 proband-parent trios of Bulgarian origin: 266 had a schizophrenic proband, 103 had a bipolar proband and 44 had a schizoaffective proband. The results were analysed using the extended transmission disequilibrium test. Possible effects of different alleles on certain clinical variables were examined by correlation analysis. Three alleles were detected: STin2.9, STin2.10 and STin2.12. None of the three diagnostic samples showed preferential transmission of alleles that reached conventional levels of statistical significance. We could not confirm previous results that STin2.12 allele increases susceptibility to bipolar disorder type I. The rare STin2.9 showed a non-significant trend for preferential transmission in the sample as a whole: 18 transmitted versus 11 non-transmitted (P = 0.2). The VNTR polymorphism in the 5-HTT gene does not appear to be a major risk factor for increasing susceptibility to major psychiatric disorders.

Age-related changes of the dentate nuclei in normal adults as revealed by 3D fast low angle shot (FLASH) echo sequence magnetic resonance imaging.

The aim of the present study was to investigate age-related changes in iron-deposition in dentate nuclei using iron-induced susceptibility effects in magnetic resonance imaging. MR images from 74 healthy subjects (age range 20-68 years) were obtained using a three-dimensional (3D) T1-weighted fast low angle shot (FLASH) echo sequence. Signal intensities of the dentate nuclei and cerebellar white matter were bilaterally measured independently by three blinded investigators. The signal intensities of dentate nuclei were intraindividually normalised to the corresponding signal intensities of the cerebellar white matter of corresponding slices. Mean normalised signal intensities were correlated with age and compared between different age decades and gender. Intraclass correlation coefficients were high (dentate nuclei: 0.98, cerebellar white matter: 0.75) indicating sufficient interrater reliabilities for the determination of signal intensities. Bland-Altman analysis confirmed this finding. The normalised mean signal intensity of the dentate nuclei correlated inversely with age (r = -0.462, p < 0.0001). Comparison of age decades revealed that significant decreases took place between the third and fourth decade and to a lesser degree between the fourth and fifth decade. Moreover, variability of normalised mean signal intensities of the dentate nuclei increased significantly with age (r = 0.964, p = 0.008). There were no differences of the normalised mean signal intensities between genders. The present study revealed an age-dependent decrease of signal intensities in dentate nuclei most likely reflecting an age-dependent increase in dentate iron concentration. These age-dependent changes have to be taken into account in interpretation of disease related MR changes of cerebellar nuclei in patients with degenerative or acquired cerebellar ataxia.

Cerebellar activation during leg withdrawal reflex conditioning: an fMRI study.

OBJECTIVE: The aim of the present study was to examine cerebellar areas related to conditioning of the nociceptive leg withdrawal reflex using event-related functional magnetic resonance imaging (fMRI). Because of the aversive nature of the unconditioned stimulus effects of accompanying fear conditioning were expected. METHODS: In 20 healthy adult subjects leg withdrawal reflex conditioning was performed using a standard delay protocol during MR-scanning. Electromyographic recordings from the anterior tibial and biceps femoris muscles were used to quantify conditioned responses. Fear-related changes of heart rate were assessed. RESULTS: In the group of all subjects a significant increase of cerebellar activation was found in the anterior and posterior vermis. In the group of subjects (n=9) who showed conditioned leg withdrawal responses cerebellar activation was more pronounced in parts of the anterior vermis, which correspond to the known leg representation. In the group of subjects (n=11) who did not develop conditioned responses cerebellar activation was more pronounced in the posterolateral hemispheres. Changes of heart rate, however, did not significantly differ between groups. CONCLUSIONS: Results suggest that areas within the anterior vermis are involved in conditioning of the leg withdrawal response. The present results, however, do not allow to differentiate between motor performance, learning or timing-related processes. Areas in the posterior vermis and cerebellar hemispheres may be related to concomitant fear conditioning. SIGNIFICANCE: Results of the present event-related fMRI study suggest involvement of the human cerebellum in conditioning of the nociceptive leg withdrawal response.