Risk factors for difficult thyroidectomy and postoperative morbidity do not match: retrospective study from an endocrine surgery academic referral centre.

Hyperthyroidism, goiter and thyroiditis have been associated with complex thyroidectomy. Difficult thyroidectomies may implicate longer operating times and higher complication rates, while literature on quantification and prediction of difficulty in thyroidectomy is scant. We aim at assessing the impact of preoperative and intraoperative factors on the technical difficulty of total thyroidectomy (TT) and on the incidence of postoperative complications. We conducted a retrospective study on 197 TT from 343 thyroidectomies performed with intraoperative neuromonitoring between October 2019 and June 2022 (excluding lobectomies, nodal dissection, extra-thyroidal procedures). Operating time (surrogate of TT difficulty), postoperative hypocalcaemia, recurrent laryngeal nerve palsy and postoperative bleeding were assessed in relation to pre- and intraoperative characteristics. Vocal fold palsy(VFP) was defined as recovering < 12 months postoperatively. There were 87 thyroid cancers and 110 multinodular goiters (21 hyperfunctioning, 51 mediastinal). Median operating time was 136 min (range 51-310). Within 17.4 months overall median follow-up we recorded two transient VFPs and 12% symptomatic transient hypocalcaemia. At univariable analysis male sex (p = 0.005), BMI (p < 0.001), thyroiditis (p < 0.05), hypervascular goiter (p = 0.003) and thyroid adhesions to surrounding anatomical structures (p < 0.001) were associated with longer operating time. At multivariable analysis male male sex (p = 0.01), obesity (p = 0.001) and thyroid adhesions (p = 0.008) were factors for prolonged operating time. Above-normal anti-thyroid peroxidase antibodies correlated to transient symptomatic hypocalcemia (p < 0.001). Risk factors for complex TT were identified and did not correlate with morbidity rates. Results from this study may help optimizing operating room schedule and inform case selection criteria for training programs in thyroid surgery. Further research is required to confirm these findings.

Indocyanine Green Fluorescence Angiography Can Guide Intraoperative Localization During Parathyroid Surgery.

BACKGROUND AND AIMS: Intraoperative localization of pathologic parathyroid glands is of major importance for the hyperparathyroidism treatment. Based on the small size and the anatomic variability, the localization can be very challenging. The current practice is to compare preoperative ultrasonography with Technetium-99m sestamibi scintigraphy (MIBI) and plan the resection accordingly. In this study, we implemented indocyanine green angiography for the intraoperative localization of parathyroid glands. MATERIALS AND METHODS: This is a retrospective analysis of 37 patients with primary, secondary, or tertiary hyperparathyroidism who were operated using indocyanine green angiography for the intraoperative localization of pathological parathyroid glands. An indocyanine green solution of 2.5 mg was were intravenously administered for parathyroid gland visualization. Different fluorescence scores were correlated with changes in postoperative parathyroid hormone levels. RESULTS: Patients were divided into two groups depending on the presence of uniglandular or multiglandular disease. Sixty-four lesions were resected, and the final histopathologic analysis confirmed the parathyroid origin in 62 of them (96.8%). None of the patients with uniglandular disease developed postoperative hypoparathyroidism, whereas three patients in the multiglandular group developed temporary hypoparathyroidism symptoms. Indocyanine green imaging had higher sensitivity for the intraoperative detection of parathyroid glands compared with ultrasonography and MIBI (p < 0.001). CONCLUSION: Indocyanine green angiography indicated high sensitivity for the intraoperative identification of pathologic parathyroid glands leading to a resection rate of 95.16%. The modality was useful, especially in cases of revisional surgery or ectopic parathyroid glands. Randomized trials have already proven the value of indocyanine green imaging in predicting postoperative hypocalcemia. Our results support the regular use of this method during parathyroid surgery.

Real-time algorithm for Poissonian noise reduction in low-dose fluoroscopy: performance evaluation.

BACKGROUND: Quantum noise intrinsically limits the quality of fluoroscopic images. The lower is the X-ray dose the higher is the noise. Fluoroscopy video processing can enhance image quality and allows further patient's dose lowering. This study aims to assess the performances achieved by a Noise Variance Conditioned Average (NVCA) spatio-temporal filter for real-time denoising of fluoroscopic sequences. The filter is specifically designed for quantum noise suppression and edge preservation. It is an average filter that excludes neighborhood pixel values exceeding noise statistic limits, by means of a threshold which depends on the local noise standard deviation, to preserve the image spatial resolution. The performances were evaluated in terms of contrast-to-noise-ratio (CNR) increment, image blurring (full width of the half maximum of the line spread function) and computational time. The NVCA filter performances were compared to those achieved by simple moving average filters and the state-of-the-art video denoising block matching-4D (VBM4D) algorithm. The influence of the NVCA filter size and threshold on the final image quality was evaluated too. RESULTS: For NVCA filter mask size of 5 × 5 × 5 pixels (the third dimension represents the temporal extent of the filter) and a threshold level equal to 2 times the local noise standard deviation, the NVCA filter achieved a 10% increase of the CNR with respect to the unfiltered sequence, while the VBM4D achieved a 14% increase. In the case of NVCA, the edge blurring did not depend on the speed of the moving objects; on the other hand, the spatial resolution worsened of about 2.2 times by doubling the objects speed with VBM4D. The NVCA mask size and the local noise-threshold level are critical for final image quality. The computational time of the NVCA filter was found to be just few percentages of that required for the VBM4D filter. CONCLUSIONS: The NVCA filter obtained a better image quality compared to simple moving average filters, and a lower but comparable quality when compared with the VBM4D filter. The NVCA filter showed to preserve edge sharpness, in particular in the case of moving objects (performing even better than VBM4D). The simplicity of the NVCA filter and its low computational burden make this filter suitable for real-time video processing and its hardware implementation is ready to be included in future fluoroscopy devices, offering further lowering of patient's X-ray dose.

Surgical management of the pancreatic stump following pancreato-duodenectomy.

Pancreato-duodenectomy (PD) is the treatment of choice for periampullary tumors, and currently, indications have been extended to benign disease, including symptomatic chronic pancreatitis, paraduodenal pancreatitis, and benign periampullary tumors that are not amenable to conservative surgery. In spite of a significant decrease in mortality in high volume centers over the last three decades (from>20% in the 1980s to<5% today), morbidity remains high, ranging from 30% to 50%. The most common complications are related to the pancreatic remnant, such as postoperative pancreatic fistula, anastomotic dehiscence, abscess, and hemorrhage, and are among the highest of all surgical complications following intra-abdominal gastro-intestinal anastomoses. Moreover, pancreatico-enteric anastomotic breakdown remains a life-threatening complication. For these reasons, the management of the pancreatic stump following resection is still one of the most hotly debated issues in digestive surgery; more than 80 different methods of pancreatico-enteric reconstructions having been described, and no gold standard has yet been defined. In this review, we analyzed the current trends in the surgical management of the pancreatic remnant after PD.

Floating left innominate vein neoplastic thrombus: a rare case of mediastinal extension of follicular thyroid carcinoma.

OBJECTIVE: We report a case of advanced follicular thyroid cancer with innominate vein involvement. To our knowledge, this seems to be the first case treated in emergency surgery, reported in literature. METHOD: A 59-year-old woman with a five-year history of a large and mainly right-sided cervical mass presented with dyspnea, unilateral arm swelling, facial flushing, and venous congestion. An emergency computed tomography scan revealed a thyroid mass extending into the upper mediastinum with displacement and compression of the right jugular vein and carotid artery and apparent adherence to the superior vena cava and left innominate vein. RESULTS: An emergency total thyroidectomy was performed by means of a sternotomy. The lower portion of the retrosternal goiter projected directly into the left innominate vein, with tumor floating in its lumen. Removal of the neoplastic thrombus was performed, through an incision in the vein, en bloc with the thyroid mass. Both goiter and thrombus were completely replaced by follicular carcinoma. CONCLUSIONS: Accurate preoperative assessment through contrast-enhanced computed tomography is strongly suggested in the presence of enlarged thyroid gland extending into the mediastinum whenever angioinvasion is suspected. This could prevent blinded maneuvers such as digital externalization of the thoracic component of the gland, which can be fatal in cases of cervico-mediastinal goiter extending into great cervical or mediastinal veins.

Recurrent laryngeal nerve palsy and substernal goiter. An Italian multicenter study.

The aim of this retrospective multicenter study was to verify whether the substernal goiter and the type of surgical access could be risk factors for recurrent laryngeal nerve palsy during total thyroidectomy. Between 1999-2008, 14,993 patients underwent total thyroidectomy. Patients were divided into three groups: group A (control; n=14.200, 94.7%), cervical goiters treated through collar incision; group B (n=743, 5.0%) substernal goiters treated by cervical approach; group C (n=50, 0.3%) in which a manubriotomy was performed. Transient and permanent unilateral palsy occurred significantly more frequently in B+C vs. A (P≤.001) and in B vs. A (P≤.001). Transient bilateral palsy was significantly more frequent in B+C vs. A (P≤.043) and in C vs. A (P≤.016). Permanent bilateral palsy was significantly more frequent in B+C vs. A (P≤.041), and in B vs. A (P≤.037). Extension of the goiter into the mediastinum was associated to increased risk of recurrent nerve palsy during total thyroidectomy.

X-Y aneuploidy rate in sperm of two "minor" breeds of cattle (Bos taurus) by using dual color fluorescent in situ hybridization (FISH).

The present study reports on the frequency of X-Y aneuploidy in the sperm population of two minor cattle breeds reared in Italy, namely Modicana and Agerolese, which are listed in the "Anagraphic Register of autochthonous cattle populations with limited distribution". More than 50 000 sperm nuclei from 11 subjects (5 and 6, respectively for each breed) have been analyzed by the fluorescent in situ hybridization with the Xcen and Y-chromosome specific painting probes. The fraction of X- and Y-bearing sperm was close to the 1:1 ratio in the Modicana breed, whereas in the Agerolese the Y-fraction was significantly higher (P < 0.002) compared to the X-counterpart. The mean rates of X-Y aneuploidy were 0.510 and 0.466%, respectively, in the two breeds; no significant differences were found among individual bulls within each breed. Average frequencies of disomic and diploid sperm were 0.425 and 0.085% in the former and 0.380 and 0.086% in the latter. In both breeds, (a) disomy was significantly more frequent than diploidy (P < 0.01), (b) YY disomy was significantly (P < 0.001) more frequent than XY or XX; (c) MI errors (XY disomy) were significantly (P < 0.01) less represented than MII (XX + YY disomy). Compared to the dairy (Italian Friesian and Brown) and meat (Podolian and Maremmana) breeds previously analyzed, the "minor" breeds investigated in the present study showed a significantly (P < 0.002) higher rate of X-Y aneuploidy (0.486 vs. 0.159 and 0.190%, respectively). Considering all the breeds analyzed -so far- and assuming no significant interchromosomal effect, the baseline level of aneuploidy in the sperm population of the species Bos taurus was estimated as 5.19%. Establishing the baseline level of aneuploidy in the sperm population of the various livestock species/breeds engaged in animal production could reveal useful for monitoring future trends of their reproductive health, especially in relation to management errors and/or environmental hazards.

Chromosome fragility in river buffalo cows exposed to dioxins.

Fifty river buffalo (Bubalus bubalis, 2n = 50) cows reared in two different provinces of Campania (southern Italy) underwent cytogenetic investigations to ascertain possible differences in their chromosome stability. One group (Caserta province) was under legal sequestration due to the presence in the milk mass of higher mean values of dioxins [21.79 pg/g of fat as sum of polychloro-dibenzo-dioxins (PCDDs), polychloro-dibenzo-furans (PCDFs) and dioxin-like polychlorobiphenyls (DL-PCBs)] than both those permitted (6.0 pg/g of fat as WHO-TEQ) and those (1.3 pg/g of fat as WHO-TEQ) observed in the control group raised in Salerno province. Two types of peripheral blood cell cultures were performed: without (normal cultures for the chromosome abnormality (CA) test: chromatid breaks, chromosome breaks, fragments) and with the addition of BrdU for the sister chromatid exchange (SCE) test). The CA test revealed a significantly (P < 0.01) higher chromosome fragility in the exposed cows compared to the control. Indeed, mean values of CA/cell were 1.26 ± 1.15 in exposed cows and 0.37 ± 0.71 in the control. Mean SCE was higher in exposed cows (8.50 ± 3.35) than that (8.29 ± 3.51) found in the control but the difference was not significant. Comparison within the same group of cows at first (FL) and multiple (ML) lactations revealed significantly (P < 0.01) higher mean values of CA/cell in exposed ML-cows vs FL-cows while no statistical differences were found between ML-cows and FL-cows in the control farm. By contrast, significantly (P < 0.01) higher mean values of SCE were found in both groups of FL-cows versus ML-cows. Comparisons with other previous studied species (sheep and cattle) were also performed.

Disorders of sexual development and abnormal early development in domestic food-producing mammals: the role of chromosome abnormalities, environment and stress factors.

The management of disorders of sexual development (DSD) in humans and domestic animals has been the subject of intense interest for decades. The association between abnormal chromosome constitutions and DSDs in domestic animals has been recorded since the beginnings of conventional cytogenetic analysis. Deviated karyotypes consisting of abnormal sex chromosome sets and/or the coexistence of cells with different sex chromosome constitutions in an individual seem to be the main causes of anomalies of sex determination and sex differentiation. In recent years, a growing interest has developed around the environmental insults, such as endocrine-disrupting compounds (EDC) and heat stressors, which affect fertility, early embryonic development and, in some instances, directly the sex ratio and/or the development of 1 specific sex versus the other. A variety of chemical compounds present in the environment at low doses has been shown to have major effects on the reproductive functions in human and domestic animals following prolonged exposure. In this review, we present an overview of congenital/chromosomal factors that are responsible for the DSDs and link them and the lack of proper embryonic development to environmental factors that are becoming a major global concern.

Similar rates of chromosomal aberrant secondary oocytes in two indigenous cattle (Bos taurus) breeds as determined by dual-color FISH.

In vitro-matured metaphase II (MII) oocytes with corresponding first polar bodies (I pb) from two indigenous cattle (Bos taurus) breeds have been investigated to provide specific data upon the incidence of aneuploidy. A total of 165 and 140 in vitro-matured MII oocytes of the Podolian (PO) and Maremmana (MA) breeds, respectively, were analyzed by fluorescence in situ hybridization using Xcen and five chromosome-specific painting probes. Oocytes with unreduced chromosome number were 13.3% and 6.4% in the two breeds, respectively, averaging 10.2%. In the PO, out of 100 MII oocytes + I pb analyzed, two oocytes were nullisomic for chromosome 5 (2.0%) and one disomic for the same chromosome (1.0%). In the MA, out of 100 MII oocytes + I pb, one oocyte was found nullisomic for chromosome 5 (1.0%) and one was disomic for the X chromosome (1.0%). Out of 200 MII oocytes + I pb, the mean rate of aneuploidy (nullisomy + disomy) for the two chromosomes scored was 2.5%, of which 1.5% was due to nullisomy and 1.0% due to disomy. By averaging these data with those previously reported on dairy cattle, the overall incidence of aneuploidy in cattle, as a species, was 2.25%, of which 1.25% was due to nullisomy and 1.0% due to disomy. The results so far achieved indicate similar rates of aneuploidy among the four cattle breeds investigated. Interspecific comparison between cattle (Xcen-5 probes) and pig (Sus scrofa domestica) (1-10 probes) also reveal similar rates. Further studies are needed that use more probes to investigate the interchromosomal effect. Establishing a baseline level of aneuploidy for each species/breed could also be useful for improving the in vitro production of embryos destined to the embryo transfer industry as well as for monitoring future trends of the reproductive health of domestic animals in relation to management errors and/or environmental hazards.

Incidence of X-Y aneuploidy in sperm of two indigenous cattle breeds by using dual color fluorescent in situ hybridization (FISH).

The present study reports on the incidence of X-Y aneuploidy in the sperm population of two indigenous cattle breeds reared in Italy for beef purposes, the Podolian and Maremmana. Totally, more than 50 000 sperm nuclei from 10 subjects (5 from each breed) have been fluorescent in situ hybridization (FISH) analyzed by using Xcen- and Y-chromosome-specific painting probes. In both breeds, the fraction of Y-bearing sperm was significantly higher (P < 0.01) compared with the X-counterpart. The rates of X-Y aneuploidy were 0.180% and 0.200%, respectively, in the Podolian and Maremmana. No significant interindividual differences were found. Average frequencies of disomic and diploid sperm were 0.149% and 0.031% in the former and 0.098% and 0.102% in the latter. Significant differences (P < 0.05) were found among the XX-XY and YY-disomy classes in both breeds, while diploidy classes were uniformly represented. In the Podolian breed, disomies were more frequent than diploidies (P < 0.05), whereas in the Maremmana they showed similar frequencies. In both breeds disomies arising from errors in meiosis I (X-Y disomies) were more represented than those arising in meiosis II (XX and YY), while this difference was not detected for diploidies. The present study provides specific information on the incidence of X-Y sperm aneuploidy in two indigenous breeds of cattle, in order to establish a breed-specific 'aneuploidy data-base' that could be used as reference for genetic improvement and future monitoring of the reproductive health of the breed.

A new and unusual reciprocal translocation in cattle: rcp(11;25)(q11;q14-21).

A new and unusual reciprocal translocation was detected in a heifer of the Agerolese cattle breed during a routine cytogenetic screening carried out on 13 animals (2 males and 11 females) kept at the ConSDABI Conservation Center in Benevento (Southern Italy). The 13 animals investigated had a normal karyotype except for a 1-year-old female, which carried one autosome smaller than the smallest normal bovine autosomes. This small autosome showed very little C-banding in comparison to the other autosomes, while another medium-sized autosome showed 2 distinct and prominent C-bands. RBA-banding and karyotype analysis revealed that these 2 chromosomes were the result of a reciprocal translocation between chromosomes 11 and 25. FISH analysis with BAC142G06 mapping to the proximal (subcentromeric) region of both BTA25 and der11, BAC513H08 (ELN) mapping to BTA25q22dist and der25, and BAC533C11 mapping to the proximal region of BTA11 and der11 confirmed the localization of the breakpoints on band q11 (centromere) of chromosome 11 and q14-21 of chromosome 25. Ag-NOR and sequential RBA/Ag-NOR techniques detected the presence of NORs on both BTA11 and BTA25 and both der11 and der25. To our knowledge, this is the first report of a reciprocal translocation event in cattle with the breakpoint located in the centromeric region.

Extended cytogenetic maps of sheep chromosome 1 and their cattle and river buffalo homoeologues: comparison with the OAR1 RH map and human chromosomes 2, 3, 21 and 1q.

Cytogenetic maps are useful tools for several applications, such as the physical anchoring of linkage and RH maps or genome sequence contigs to specific chromosome regions or the analysis of chromosome rearrangements. Recently, a detailed RH map was reported in OAR1. In the present study, we selected 38 markers equally distributed in this RH map for identification of ovine genomic DNA clones within the ovine BAC library CHORI-243 using the virtual sheep genome browser and performed FISH mapping for both comparison of OAR1 and homoeologous chromosomes BBU1q-BBU6 and BTA1-BTA3 and considerably extending the cytogenetic maps of the involved species-specific chromosomes. Comparison of the resulting maps with human-identified homology with HSA2q, HSA3, HSA21 and HSA1q reveals complex chromosome rearrangements differentiating human and bovid chromosomes. In addition, we identified 2 new small human segments from HSA2q and HSA3q conserved in the telomeric regions of OAR1p and homoeologous chromosome regions of BTA3 and BBU6, and OAR1q, respectively. Evaluation of the present OAR1 cytogenetic map and the OAR1 RH map supports previous RH assignments with 2 main exceptions. The 2 loci BMS4011 and CL638002 occupy inverted positions in these 2 maps.

A rare case of centric fission and fusion in a river buffalo (Bubalus bubalis, 2n = 50) cow with reduced fertility.

A 5-year-old river buffalo cow underwent cytogenetic investigation since it had only one male offspring, apparently with normal body constitution, which died one month after birth. The female carrier had normal body conformation and internal sex adducts, as revealed by rectal palpation performed by a specialist veterinary practitioner. The cow was found to carry a complex and rare chromosome abnormality. Indeed, a centric fission of one river buffalo (BBU) chromosome 1 with a subsequent (or simultaneous) centric fusion of BBU1p with BBU23 was revealed by both RBA-banding and specific molecular markers of BBU1p (DEFB1) and BBU23 (ACTA2). CBA-banding revealed a pale, very small C-band in the der1 (BBU1q) and a prominent C-band on the new biarmed chromosome originated by rob(1p;23). Both telomeric probes and AgNOR staining confirmed the Robertsonian translocation (rob), both FITC-signals and the NORs (BBU23) being telomerically located. Furthermore, telomeric signals on der1 (BBU1q) indicate that these 2 chromosomal events may be the result of a reciprocal translocation which occurred between BBU1 and BBU23.

Chromosome fragility in dairy cows exposed to dioxins and dioxin-like PCBs.

In this study, we compared cross-bred dairy cows in the Susa Valley (Piedmont, northern Italy), reared either near a high-temperature steel production plant (Farms A and B) or in an industry-free area (control). Exposed cows (n = 36) were selected based on mean bulk milk toxic equivalent values of polychlorodibenzodioxins (PCDDs) and dioxin-like (DL) polychlorobiphenyls (PCBs) and polychlorodibenzofurans (PCDFs) equal to 18.56 pg/g fat and 8.56 pg/g of fat in dairy cows from Farms A and B, respectively, exceeding both those permitted by the legislation in force (6 pg/g fat PCDDs and DL-PCDFs/PCBs), and those measured in dairy cows (n = 19) of the farm used as control (1.75 pg/g of fat PCDDs and DL-PCDFs/PCBs). Two types of peripheral blood cell cultures were performed: without (normal cultures for the chromosome abnormality (CA)-test: gaps, chromatid breaks, chromosome breaks and fragments) and with addition of bromodeoxyuridine [for the sister chromatid exchange (SCE)-test]. Both tests revealed a significant (P ≤ 0.05) higher chromosome fragility in the exposed cattle compared to controls: CA/cell mean values (without gaps) were 0.65 ± 0.91, 0.51 ± 0.81 and 0.13 ± 0.39 in Farms A, B and controls, respectively, while SCE/cell mean values were 7.00 ± 2.88, 6.39 ± 2.80 and 5.29 ± 2.51. Although the role of other pollutants (e.g. heavy metals) in the genesis of the recorded chromosome alterations cannot be ruled out, our results confirm the findings of previous research into dioxin-exposed sheep.

Cytogenetic and genetic studies in a hypospadic horse (Equus caballus, 2n = 64).

A 4-year-old male horse of Friesian breed with normal body conformation, development and libido, and showing an evident ventral penis deviation with hypospadias, underwent both cytogenetic and genetic investigation. Although the karyotype showed normal male arrangement (2n = 64,XY), one telomere of horse (ECA) chromosome 1 was shorter than both the other one and those of a normal horse (control), as revealed by CBA- and RBA-banding, and by Ag-NOR and FISH-mapping techniques using telomere PNA probes. Genetic investigation of the SRY and MAMLD1 coding sequences revealed a normal SRY sequence and a mutation in the MAMLD1 gene sequence: a homozygous change (C>A) was found, leading to the synthesis of an isoleucine, instead of a leucine. Although it is difficult to find a strict correlation between hypospadias and the genetic defects revealed by this investigation, this study is the first to be performed in a hypospadic horse using both cytogenetic and genetic investigation.

Chromosomal assignment of R-spondin genes in the donkey (Equus asinus, 2n = 62).

R-spondins constitute a recently discovered small family of growth factors, and the evidence of their role in several developmental pathways is growing fast. In this work we describe the chromosomal location of the four RSPO genes in the donkey. Using horse BACs, we localized RSPO1 on EAS 5q23, RSPO2 on EAS 12q13, RSPO3 on EAS 24q26, and RSPO4 on EAS 15p13. Moreover, RSPO2, RSPO3, and RSPO4 are the first genes mapped on donkey chromosomes 12, 24, and 15, respectively.

Genomic characterization and chromosomal mapping of 5 river buffalo skeletal muscle differentiation master genes.

River buffalo (Bubalus bubalis, 2n = 50, BBU) is a species of economic relevance in a number of countries. This species shows a very peculiar biology and a great capacity for environmental adaptation. There has been an increasing economic interest as well as a growing demand for a more detailed knowledge of molecular features in this species. From this perspective we report a genomic, transcriptional and cytogenetic analysis of 5 master genes involved in skeletal muscle development. Of these 5 genes, MYOD1, MYF5, MYF6 and MYOG belong to the basic helix-loop helix protein family while MSTN belongs to the TNF-B protein family. In mammals, these genes are involved in the early stages of skeletal muscle differentiation, development and regeneration. These pivotal biological functions are finely regulated in a tissue- and temporal-specific manner. We used a comparative genomic approach to obtain the buffalo specific sequences of MYOD1 and MYF6. The nucleotide sequence similarity and the protein domain conservation of the newly obtained sequences are analysed with respect to bovine and other mammalian species showing sequence similarity. The presence of a polymorphism in MYOD1 coding sequence is described and its possible effect discussed. Using a quantitative PCR approach, we compared the level of the 5 transcripts in adult and fetal muscle. These genes were physically localised on river buffalo R-banded chromosomes by FISH using bovine genomic BAC-clones. Here, we present a genomic and cytogenetic analysis which could offer a background to better characterise the buffalo genes involved in muscle function and which may be responsible for buffalo-specific meat features.

Frequency of aneuploidy in in vitro-matured MII oocytes and corresponding first polar bodies in two dairy cattle (Bos taurus) breeds as determined by dual-color fluorescent in situ hybridization.

The current study was undertaken to investigate the aneuploidy rates in in vitro-matured meiosis II (MII) oocytes and corresponding first polar bodies in two dairy cattle (Bos taurus) breeds by using dual-color fluorescent in situ hybridization (FISH). A total of 159 and 144 in vitro-matured MII oocytes of the Italian Friesian and Italian Brown breeds, respectively, were obtained according to the standard methods and analyzed by FISH using "Xcen" and "5" chromosome-specific painting probes, produced by chromosome microdissection and Degenerate Oligonucleotide Primer- Polymerase Chain Reaction (DOP-PCR). Oocytes with unreduced chromosome number were 10.1% and 16.7% in the two breeds, respectively. To avoid bias due to possible artifacts, the aneuploidy rates were determined by analyzing only oocytes with the corresponding polar bodies. In the Italian Friesian, 100 of 143 (69.9%) secondary MII oocytes showed clear MII plates with corresponding first polar bodies and were scored for aneuploidy detection; one oocyte was "nullisomic" for chromosome X (1.0%) and one "disomic" for chromosome 5 (1.0%). In the Italian Brown, 100 of 120 (83.3%) MII oocytes with corresponding first polar bodies were analyzed; one oocyte was nullisomic (1.0%) and one was disomic (1.0%), both for chromosome 5. Totally, 303 oocytes were analyzed, 40 of which showed an unreduced chromosome complement (13.2%); of 200 MII oocytes with the corresponding first polar bodies, the aneuploidy rate (nullisomy+disomy) for the two chromosomes scored was 2%. Assuming that each chromosome is equally involved in aneuploidy, it results that in cattle oocytes matured in vitro, at least 30% of the oocytes (1x30 haploid chromosomes) should be aneuploid. Premature separation of sister chromatids (PSSC) was also observed in 2% of the oocytes in the Italian Friesian breed involving chromosome 5 and in 1% of the Italian Brown breed involving the X chromosome. Estimation of the "baseline" level of aneuploidy in the in vitro-matured oocytes of the various domestic animal species and breeds is, to our opinion, a useful reference for improving the in vitro production of embryos as well as for monitoring future trends of the reproductive health of the species/breeds engaged in zootechnical productions, especially in relation to management errors and environmental hazards.

Molecular cytogenetics and gene mapping in sheep (Ovis aries, 2n = 54).

The development of a completely annotated sheep genome sequence is a key need for understanding the phylogenetic relationships and genetic diversity among the many different sheep breeds worldwide and for identifying genes controlling economically and physiologically important traits. The ovine genome sequence assembly will be crucial for developing optimized breeding programs based on highly productive, healthy sheep phenotypes that are adapted to modern breeding and production conditions. Scientists and breeders around the globe have been contributing to this goal by generating genomic and cDNA libraries, performing genome-wide and trait-associated analyses of polymorphism, expression analysis, genome sequencing, and by developing virtual and physical comparative maps. The International Sheep Genomics Consortium (ISGC), an informal network of sheep genomics researchers, is playing a major role in coordinating many of these activities. In addition to serving as an essential tool for monitoring chromosome abnormalities in specific sheep populations, ovine molecular cytogenetics provides physical anchors which link and order genome regions, such as sequence contigs, genes and polymorphic DNA markers to ovine chromosomes. Likewise, molecular cytogenetics can contribute to the process of defining evolutionary breakpoints between related species. The selective expansion of the sheep cytogenetic map, using loci to connect maps and identify chromosome bands, can substantially contribute to improving the quality of the annotated sheep genome sequence and will also accelerate its assembly. Furthermore, identifying major morphological chromosome anomalies and micro-rearrangements, such as gene duplications or deletions, that might occur between different sheep breeds and other Ovis species will also be important to understand the diversity of sheep chromosome structure and its implications for cross-breeding. To date, 566 loci have been assigned to specific chromosome regions in sheep and the new cytogenetic map is presented as part of this review. This review will also summarize the current cytogenomic status of the sheep genome, describe current activities in the sheep cytogenomics research sector, and will discuss the cytogenomics data in context with other major sheep genomics projects.