Molecular and Serological Detection of Bovine Coronaviruses in Marmots (Marmota marmota) in the Alpine Region.

In this study, virological surveillance focused on coronaviruses in marmots in the Alpine region in 2022, captured as part of a population control reduction program in the Livigno area. Seventy-six faecal samples were randomly collected from marmots at the time of capture and release and tested for genome detection of pan-coronavirus, pan-pestivirus, canine distemper virus, and influenza A and D virus. Nine faecal samples were positive in the Pan-CoV RT-PCR, while all were negative for the other viruses. Pan-coronavirus positives were further identified using Illumina's complete genome sequencing, which showed the highest homology with Bovine Coronavirus previously detected in roe deer in the Alps. Blood samples (n.35) were collected randomly from animals at release and tested for bovine coronavirus (BCoV) antibodies using competitive ELISA and VNT. Serological analyses revealed that 8/35 sera were positive for BCoV antibodies in both serological tests. This study provides molecular and serological evidence of the presence of BCoV in an alpine marmot population due to a likely spillover event. Marmots share areas and pastures with roe deer and other wild ruminants, and environmental transmission is a concrete possibility.

Comparative Analysis of the pIgR Gene from the Antarctic Teleost Trematomus bernacchii Reveals Distinctive Features of Cold-Adapted Notothenioidei.

The IgM and IgT classes were previously identified and characterized in the Antarctic teleost Trematomus bernacchii, a species belonging to the Perciform suborder Notothenoidei. Herein, we characterized the gene encoding the polymeric immunoglobulin receptor (pIgR) in the same species and compared it to the pIgR of multiple teleost species belonging to five perciform suborders, including 11 Antarctic and 1 non-Antarctic (Cottoperca gobio) notothenioid species, the latter living in the less-cold peri-Antarctic sea. Antarctic pIgR genes displayed particularly long introns marked by sites of transposable elements and transcription factors. Furthermore, analysis of T. bernacchii pIgR cDNA unveiled multiple amino acid substitutions unique to the Antarctic species, all introducing adaptive features, including N-glycosylation sequons. Interestingly, C. gobio shared most features with the other perciforms rather than with the cold-adapted relatives. T. bernacchii pIgR transcripts were predominantly expressed in mucosal tissues, as indicated by q-PCR and in situ hybridization analysis. These results suggest that in cold-adapted species, pIgR preserved its fundamental role in mucosal immune defense, although remarkable gene structure modifications occurred.

Mycobacterium aviumsubsp. paratuberculosis isolated from wild red deer (Cervus elaphus) in Northern Italy.

Paratuberculosis (or Johne's disease) is an infectious disease which affects mainly ruminants and it is caused by Mycobacterium avium subsp. paratuberculosis (MAP). During a culling program (years 2011-2015) aimed at controlling the red deer (Cervus elaphus) population in Stelvio National Park (Italian Alps), where paratuberculosis was already described in this species, 382 tissue samples from the Lombardy Region and 102 fecal specimens from the Autonomous Province of Bolzano were analyzed by PCR. Of these, 77 samples (20.16%) from the Lombardy area and 19 specimens (18.63%) from the Bolzano area resulted PCR positive. The cultural test was carried out on PCR positive samples (n = 96), enabling the isolation of 19 MAP field strains which were genotyped using MIRU-VNTR typing and Short Sequence repeats (SSRs). Our results suggest that all isolates share an identical VNTR profile corresponding to the INMV1 genotype. The only variation was on the locus SSR2, but the utility of this last locus has already been questioned because of its instability. Overall, these data suggest a common clonal origin and host adaptation during the diffusion of paratuberculosis in this population. Finally, this profile is the same as that which has already been described in the cattle population in Northern Italy, suggesting a possible inter-species disease transmission pattern from wildlife to domestic ruminants and vice versa.

Morphological and molecular characterization of four Sarcocystis spp., including Sarcocystis linearis n. sp., from roe deer (Capreolus capreolus) in Italy.

Fresh (frozen/thawed) muscle samples from four 2-12-year-old roe deer (Capreolus capreolus) from the Sondrio province in north-eastern Italy were examined under a dissecting microscope, and about 180 sarcocysts were isolated and identified to morphological type in wet mounts by light microscopy (LM). Seventy-seven of these sarcocysts were subsequently examined by molecular methods, comprising polymerase chain reaction (PCR) amplification and sequencing of the partial cytochrome c oxidase subunit I gene (cox1) of all isolates, as well as PCR amplification, cloning and sequencing of the complete18S ribosomal RNA (rRNA) gene of two isolates of each species found. By LM, three major sarcocyst types were recognised: spindle-shaped sarcocysts, 0.5-3 mm long, either with no clearly recognisable protrusions (thin-walled) or with finger-like protrusions (thick-walled); and slender, thread-like sarcocysts, 2-3 mm long, with hair-like protrusions. Sequencing of cox1 revealed that the sarcocysts belonged to four different species. Those with no visible protrusions either belonged to Sarcocystis gracilis (n = 24) or to a Sarcocystis taeniata-like species (n = 19), whereas those with finger- and hair-like protrusions belonged to Sarcocystis silva (n = 27) and Sarcocystis capreolicanis (n = 7), respectively. The 19 cox1 sequences of the S. taeniata-like species, comprising five haplotypes, differed from each other at 0-16 of 1038 nucleotide positions (98.5-100% identity). They differed from 25 previous cox1 sequences of S. taeniata from moose and sika deer (with 98.0-100% intraspecific identity), at 33-43 nucleotide positions (95.9-96.8% interspecific identity), and there were 20 fixed nucleotide differences between the two populations. In the phylogenetic analysis based on cox1 sequences, the two populations formed two separate monophyletic clusters. The S. taeniata-like species in roe deer was therefore considered to represent a separate species, which was named Sarcocystis linearis n. sp. At the 18S rRNA gene, however, the two species could not be clearly separated from each other. Thus, there was considerable intraspecific sequence variation in the 18S rRNA gene of S. linearis (98.1-99.9% identity between 24 sequences), which was similar both in magnitude and nature to the variation previously found in this gene of S. taeniata. The new 18S rRNA gene sequences of S. linearis shared an identity of 97.9-99.6% with those of S. taeniata (overlap between intra- and interspecific identity), and in the phylogenetic tree, sequences of the two species were interspersed. By scanning electron microscopy (SEM), the sarcocysts of S. linearis were found to possess regularly spaced, thin and narrow ribbon-like cyst wall protrusions (about 2.8-3.2 µm long, 0.3-0.4 µm wide and about 0.02-0.03 µm thick), terminating in a plate-like structure of the same thickness but with an elliptic outline (about 0.3-0.4 µm wide and 0.7-0.9 µm long). The terminal plates were connected in the middle with the band-like portion of the protrusions like the board of a seesaw (tilting board). The terminal plates of adjacent protrusions were neatly arranged in a hexagonal pattern resembling tiles on a roof. Together, they formed an outer roof-like layer facing the surrounding cytoplasm of the host cell and completely covering the band-like proximal portion of the protrusions, which overlapped and were stacked in three to four layers close to the cyst surface. The sarcocyst morphology of S. linearis was consistent with that of an unnamed Sarcocystis sp. in roe deer previously found by transmission electron microscopy in several countries, including Italy. A few sarcocysts of S. gracilis and S. silva were also examined by SEM, confirming the presence of regularly distributed, short knob-like protrusions in S. gracilis (as seen in previous SEM studies) and revealing tightly packed, erect 6-7-µm-long villus-like protrusions having regularly distributed round depressions on their surface in S. silva. The sequencing of cox1 of 7, 24 and 27 new isolates of S. capreolicanis, S. gracilis and S. silva, respectively, recovered 7, 11 and 10 new haplotypes from each of the three species and expanded our knowledge on the intraspecific sequence variation at this marker. Similarly, the study revealed a more extensive intragenomic sequence variation at the 18S rRNA gene of S. capreolicanis and S. silva than known from previous studies and confirmed a near absence of such variation in the 18S rRNA gene of S. gracilis.

Immune response of the Antarctic teleost Trematomus bernacchii to immunization with Psychrobacter sp. (TAD1).

Adult Trematomus bernacchii have been immunized intraperitoneally with heat-killed cells of the Antarctic marine bacterium Psychrobacter sp. (TAD1) up to 60 days. After immunizations and sampling at various times, fish sera were tested for specific IgM by ELISA, and different tissues (head kidney and spleen) were investigated for transcription of master genes of the acquired immune response (IgM, IgT, TRß, TRγ). Results from ELISA assays showed a time-dependent induction of specific serum anti-TAD1 IgM, and western blot analysis of TAD1 lysates probed with fish sera revealed enhanced immunoreactivity in immunized animals compared to controls. Quantitative PCR analysis of transcripts coding for IgM, IgT, TRß, TRγ was performed in T. bernacchii tissues to assess basal expression, and then on cDNAs of cells from head kidney and spleen of fish injected for 8, 24, and 72 h with inactivated TAD1. The results showed a differential basal expression of transcripts in the examined tissues, and a time-dependent strong up-regulation of IgT, TRß, TRγ genes upon in vivo stimulation with TAD1. These results represent a first in vivo study on the mounting of a specific immune response in an Antarctic teleost species.

New insights into evolution of IgT genes coming from Antarctic teleosts.

Cloning and characterization of IgT heavy chain genes were performed in the Antarctic Notothenioid teleost Trematomus bernacchii and in a non-Antarctic Notothenioid species, Bovichtus diacanthus, belonging to a phyletically basal lineage of Notothenioids. Compared to IgT from other non-Antarctic teleost species, including B. diacanthus, T. bernacchii IgT lacked most of the second constant domain but maintained only a few amino acid residues, which could be aligned to B. diacanthus CH2 domain. By analyzing several cDNA clones from a single specimen, three differently sized IgT transcript variants, named Long, Short and Shortest, were identified. Genomic analysis of T. bernacchii and B. diacanthus IgH loci revealed that, in the case of T. bernacchii, within the intron between the exons coding for the entire first and second constant domains a reminiscence of the ancestral second exon was present. The Long and Short variants were found to be encoded by indel alleles, whereas the Shortest variant was generated by alternative splicing that led to the CH2 exonic remnant skipping. Through comparison between genomic and cDNA sequences we hypothesized the presence of three different copies of the IgT heavy chain gene, one of which being considered the functional gene since the corresponding transcripts were identified. Moreover, either Long or Short exonic variants were found to be used in IgT heavy chain membrane form in an unbiased manner, as seen for the secretory form. Phylogenetic analysis was performed on the constant region from all teleost IgT available to date, including IgT from another Antarctic Notothenioid species, Notothenia coriiceps, identified by searching the transcriptome. The loss of almost an entire domain together with the conservation of some amino acids such as proline, glycine and cysteine in the CH2 domain remnant, could be interpreted as another distinctive feature of the Antarctic fish genome evolution, providing also new insights into the structural variation of teleost immunoglobulin genes.

Evolution of the complement system C3 gene in Antarctic teleosts.

Notothenioidei are typical Antarctic teleosts evolved to adapt to the very low temperatures of the Antarctic seas. Aim of the present paper is to investigate sequence and structure of C3, the third component of the complement system of the notothenioid Trematomus bernacchii and Chionodraco hamatus. We determined the complete nucleotide sequence of two C3 isoforms of T. bernacchii and a single C3 isoform of C. hamatus. These sequences were aligned against other homologous teleost sequences to check for the presence of diversifying selection. Evidence for positive selection was observed in the evolutionary lineage of Antarctic teleost C3 sequences, especially in that of C. hamatus, the most recently diverged species. Adaptive selection affected numerous amino acid positions including three residues located in the anaphylatoxin domain. In an attempt to evaluate the link between sequence variants and specific structural features, we constructed molecular models of Antarctic teleost C3s, of their proteolytic fragments C3b and C3a, and of the corresponding molecules of the phylogenetically related temperate species Epinephelus coioides, using human crystallographic structures as templates. Subsequently, we compared dynamic features of these models by molecular dynamics simulations and found that the Antarctic C3s models show higher flexibility, which likely allows for more pronounced movements of both the TED domain in C3b and the carboxyl-terminal region of C3a. As such dynamic features are associated to positively selected sites, it appears that Antarctic teleost C3 molecules positively evolved toward an increased flexibility, to cope with low kinetic energy levels of the Antarctic marine environment.

Investigation of immunoglobulins in skin of the Antarctic teleost Trematomus bernacchii.

The presence and production of IgM in the skin of the Antarctic teleost Trematomus bernacchii were investigated in this study. Immunoglobulins purified from cutaneous mucus and analysed by SDS-PAGE run under non-reducing and reducing conditions, were composed of heavy and light chains of 78 kDa and 25 kDa respectively, with a relative molecular mass of 830 kDa indicating that mucus IgM are tetramers as the serum IgM. Mature transcripts encoding the constant domains of both the secretory and membrane-bound Igµ chain were seen in T. bernacchii skin using a PCR strategy and the expression of the secretory Igµ chain in the skin was compared with that in other tissues by Real-time PCR. Cytological investigations revealed the presence of either immunoglobulins or their transcripts in occasional lymphocytes distributed close to the basal membrane. IgM once produced here, enters the filament-containing cells and is released into the mucus when these cells degenerate and detach from the epidermis. Our findings indicate that a cutaneous defence mechanism, functioning as anatomical and physiological barrier under subzero conditions, is present in this Antarctic species as an important component of the immune system.

Allelic polymorphism of Immunoglobulin heavy chain genes in the Antarctic teleost Trematomus bernacchii.

IgM represents the main immunoglobulin isotype shared by all teleost fish. However, Antarctic fish IgM possess a peculiar hinge region, which connects the CH2 and CH3 domains, not seen in any other teleost species. In the present study allelic polymorphism of IgM gene of the Antarctic teleost Trematomus benacchii was investigated. By nucleotide sequencing the entire Immunoglobulin heavy chain constant region from ten T. bernacchii individuals, 47 positions were found to be polymorphic. The largest number of polymorphic positions, accounting for 51% of the total, was found to fall within the hinge region. This region not only displayed extensive nucleotide variation, but also length diversity; in fact several sequences were one amino acid shorter as resulting from the usage of a different splice acceptor site of the CH3 exon, as demonstrated by genomic DNA analysis. The Ka/Ks ratios of the polymorphic positions showed typical values higher than 1, indicative of positive selection acting to polymorphic codons to favor amino acid replacements and maintain allelic variants.

Immune competence of the Ciona intestinalis pharynx: complement system-mediated activity.

In the tunicate Ciona intestinalis, the ciliated pharynx, which connects the external environment to a highly developed and compartmentalized gastrointestinal system, represents the natural portal of entry for a vast and diverse, potentially pathogenic microbial community. To address the role of the pharynx in immune surveillance in Ciona, we asked whether C3, the key component of the complement system, was expressed in this organ and whether the encoded protein was functionally active. We found by real-time PCR that C3, constitutively expressed in the pharynx, is up-regulated by LPS injection. Using two specific anti-CiC3 and anti-CiC3a polyclonal antibodies in immunohistochemical staining of pharynx sections, we found that the gene product was localized to hemocytes of the pharyngeal bars (identified as granular amoebocytes) and in stigmata ciliated cells. Use of the same antibodies in Western blot analysis indicated that CiC3 and its activation products CiC3b and CiC3a are present in pharynx homogenates. Our observation that the amount of the bioactive fragment CiC3a increased in the pharynx of LPS-treated animals provides the first molecular and functional evidence for complement-mediated immunological activity in the tunicate pharynx.

Evolutionary analysis of Antarctic teleost Toll-like receptor 2.

In the present study we address the investigation of TLR2 evolutionary selection in two Antarctic teleosts, Trematomus bernacchii (Nototheniidae) and Chionodraco hamatus (Channichthyidae). The nucleotide sequence of TLR2 has been determined in both species, encoding 20 leucine-rich repeats (LRRs) in the extracellular region and a classical Toll/IL-1R (TIR) domain in the intracellular region. High expression level of T. bernacchii TLR2 was found in spleen and skin. Using different methods we identified six codons that underwent Darwinian selection while 20 were found to be negatively selected. Molecular models of C. hamatus and T. bernacchii TLR2 ectodomain as well as of the TIR domain were built by Homology Modeling. Molecular Dynamics simulations were performed in water for 15 ns. The sites under positive selection were residing on the convex side of the solenoid, four out of six were in a 35-residue-long region including the central/N-terminal domain boundary: two in the external loop of LRR11 and the other two in the LRR12 loop. This region has been demonstrated to be the functional site of ligand interaction in human TLR2 structure. Antarctic TLR2 models showed more flexibility than TLR2 from the temperate species Gasterosteus aculeatus. These results suggest that the selective pressure has shaped TLR2 molecule in such a way that increased its activity under the peculiar Antarctic environmental conditions.

Immunoglobulin from Antarctic fish species of Rajidae family.

Immunoglobulins (Ig) of Chondroichthyes have been extensively studied in sharks; in contrast, in skates investigations on Ig remain scarce and fragmentary despite the high occurrence of skates in all of the major oceans of the world. To focus on Rajidae Igµ, the most abundant heavy chain isotype, we have chosen the Antarctic species Bathyraja eatonii, Bathyraja albomaculata, Bathyraja brachyurops, and Amblyraja georgiana which live at high latitudes in the Southern Ocean, and at very low temperatures. We prepared mRNA from the spleen of individuals of each species and performed RT-PCR experiments using two oligonucleotides designed on the alignment of various elasmobranch Igµ heavy chain sequences available in GenBank. The PCR products, about 1400-nt long, were cloned and sequenced. Nucleotide sequence identities calculated for the constant region domains ranged from 88.5% to 97.5% between species, and from 91.1% to 99.7% within species. In a distance tree, including also Raja erinacea sequences, two major branches were obtained, one containing Arhynchobatinae sequences, the other one Rajinae sequences. Four presumptive D gene segments were identified in the region of the VH/D/JH recombination; two different D segments were often found in the same sequence. Moreover, 5-15 genomic fragments of different lengths, carrying the gene locus encoding Igµ chain were revealed by Southern blotting analysis. B. eatonii amino acid sequences were analyzed for the positional diversity by Shannon entropy analysis, showing CH4 as the most conserved domain, and CH3 as the most variable one. B. eatonii CDR3 region length varied between 11 and 15 amino acid residues; the mean length (13.4 aa) was greater than that of Leucoraja eglanteria sequences (7.7 aa). An alignment of representative sequences of Antarctic species and R. erinacea showed that more cysteine residues not involved in the intradomain disulfide bridges were present in Antarctic species.

A role for variable region-containing chitin-binding proteins (VCBPs) in host gut-bacteria interactions.

A number of different classes of molecules function as structural matrices for effecting innate and adaptive immunity. The most extensively characterized mediators of adaptive immunity are the immunoglobulins and T-cell antigen receptors found in jawed vertebrates. In both classes of molecules, unique receptor specificity is effected through somatic variation in the variable (V) structural domain. V region-containing chitin-binding proteins (VCBPs) consist of two tandem Ig V domains as well as a chitin-binding domain. VCBPs are encoded at four loci (i.e., VCBPA-VCBPD) in Ciona, a urochordate, and are expressed by distinct epithelial cells of the stomach and intestine, as well as by granular amoebocytes present in the lamina propria of the gut and in circulating blood. VCBPs are secreted into the gut lumen, and direct binding to bacterial surfaces can be detected by immunogold analysis. Affinity-purified native and recombinant VCBP-C, as well as a construct consisting only of the tandem V domains, enhance bacterial phagocytosis by granular amoebocytes in vitro. Various aspects of VCBP expression and function suggest an early origin for the key elements that are central to the dialogue between the immune system of the host and gut microflora.

Antarctic teleost immunoglobulins: more extreme, more interesting.

We have investigated the immunoglobulin molecule and the genes encoding it in teleosts living in the Antarctic seas at the constant temperature of -1.86 °C. The majority of Antarctic teleosts belong to the suborder Notothenioidei (Perciformes), which includes only a few non-Antarctic species. Twenty-one Antarctic and two non-Antarctic Notothenioid species were included in our studies. We sequenced immunoglobulin light chains in two species and µ heavy chains, partially or totally, in twenty species. In the case of heavy chain, genomic DNA and the cDNA encoding the secreted and the membrane form were analyzed. From one species, Trematomus bernacchii, a spleen cDNA library was constructed to evaluate the diversity of VH gene segments. T. bernacchii IgM, purified from the serum and bile, was characterized. Homology Modelling and Molecular Dynamics were used to determine the molecular structure of T. bernacchii and Chionodraco hamatus immunoglobulin domains. This paper sums up the previous results and broadens them with the addition of unpublished data.

Evolution of the Antarctic teleost immunoglobulin heavy chain gene.

Notothenioid teleosts underwent major modifications of their genome to adapt to the cooling of the Antarctic environment. In order to identify specific features of the Antarctic teleost immunoglobulin, transcripts encoding the constant region of the IgM heavy chain from 13 Antarctic and non-Antarctic notothenioid species were sequenced. The primary mRNA splicing for the membrane form was found to be atypical in the majority of Antarctic species, because it led to exclusion of two entire constant exons, and to inclusion of 39-nucleotide exons encoding an unusually long Extracellular Membrane-Proximal Domain (EMPD). Genomic DNA analysis revealed that each 39-nucleotide exon fell within a long sequence that was the reverse complement of an upstream region. Deduced amino acid sequence analysis lead to the identification of cysteine encoding codons in the 39-nucleotide exons, but not in the respective sequence counterpart, suggesting that these residues might play an important role in the folding of the EMPD.

Immunoglobulin light chain isotypes in the teleost Trematomus bernacchii.

Three immunoglobulin light chain (IgL) isotypes TrbeL1, TrbeL2, and TrbeL3 were identified in the Antarctic teleost Trematomus bernacchii by immunoscreening a cDNA expression library, and using RT-PCR, and 5' RACE. One of them was distinguished in two subisotypes TrbeL1A and TrbeL1B. Real-time PCR experiments showed that the different isotypes were expressed in similar ratios in the various tissues analyzed. Interestingly, the expression level of TrbeL1A isotype was very high in all tissues. Molecular models of the CH1-CL domain pairings were built and minimized for the different isotypes. Several differences were identified in the superimposable structures mainly in the loops. In addition, the isotype-specific residues determined a different distribution of the charges on the external CL domain surface. Phylogenetic trees of 43 isotype representative sequences of CL domain from teleost species, built by different methods, indicated that all teleost light chain isotypes are distributed into three groups. Furthermore, the split of the group IgL1 into two subgroups, one of them carrying a micro-satellite DNA insertion, may have occurred in the Acanthopterygean ancestor.

Acute alcoholic hepatitis.

Acute alcoholic hepatitis (AAH) is a frequent inflammatory liver disease with high short-term mortality rate. In this review, relationships between alcohol abuse and the epidemiology and the outcomes of AAH are discussed, as well as AAH pathogenesis. The role of endotoxins, tumor necrosis factor alpha, fibroblasts, and immune response to altered hepatocyte proteins is discussed. The need of a careful prognosis, supported by the use of Maddrey score, by the model for end-stage liver disease [Mayo end-stage liver disease (MELD)] score or by the Glasgow alcoholic hepatitis score, is outlined, as the use of the most effective drugs (glucocorticoids and anti-tumor necrosis factor alpha infliximab) is recommended only in severe AAH cases. The problems of liver transplant in severe AAH, and the need of a 6-month alcohol abstinence before transplant, are discussed, as well as the need of a careful psychologic assessment before the transplant.

MRI and muscle signal intensities in alcoholics compared with control subjects.

OBJECTIVE: To evaluate by magnetic resonance imaging (MRI) the difference in muscle signal intensities between alcoholics and control subjects. METHODS: Thirty-four healthy subjects and 46 alcohol-dependent individuals were assessed. MRI was carried out using a low-field magnet (0.2 Tesla) and a limb-dedicated coil. The presence of muscle changes was evaluated by measuring signal intensities at the medial (MG) and lateral heads of the gastrocnemius muscle by T1-/T2-weighted and gradient-echo short tau inversion recovery sequences. The mean signal intensities of the two sample groups were compared by ANCOVA with age as a covariate. In the alcohol-dependent group, correlations between signal intensities and plasma levels of muscular and hepatic enzymes, in addition to years of high-risk consumption and lifetime dose of ethanol consumed, were assessed. The mean signal intensities were also compared with the different degrees of pain by ANOVA. RESULTS: Compared with healthy subjects, the alcohol-dependent group had mean higher signal intensities in both gastrocnemius heads in all sequences. The difference in the MG in T2-weighted sequences was significant (F = 48.28, p < 0.01). A modest correlation between the years of high-risk consumption and the signal intensity was found in T2-weighted sequences in the MG (r = 0.288, p = 0.057), whereas a correlation with the lifetime dose consumed was not found. Significant correlations between signal intensities and plasma levels of muscular and hepatic enzymes were not found. There were also no significant group differences on different degrees of pain. CONCLUSION: MRI was shown to be a sensitive, well-tolerated, and inexpensive procedure capable of detecting changes in signal intensities in the muscles of alcoholics. This technique could be included among other diagnostic tools for alcoholic myopathy with further improvements and if the signal alterations can be corroborated by biopsy evidence.