Nucleolar Organizer Region Staining Research Articles

Modified silver staining of nucleolar organizer regions to improve the accuracy of image analysis

Silver staining of nucleolar organizer regions (NORs) and their subsequent quantification by image analysis are used increasingly in human pathological specimens and experimental models. Because certain conditions determined by the type of tissue and/or its fixation render AgNOR segmentation for image analysis difficult due to insufficient contrast or nonspecific silver precipitation, we propose three improvements to the original technique to overcome these difficulties. Pretreatment with 7% nitric acid produced very distinct dark brown images of AgNORs on a yellow background. The gradient of background colors allowed easy discrimination of nucleolar, nuclear and cytoplasmic structures. Seven morphometric parameters related to number, size and shape of AgNORs were evaluated quantitatively by image analysis on sections pretreated with nitric acid and on adjacent sections treated with citrate buffer in a wet autoclave according to the most widely accepted method for image analysis of AgNOR. Both methods yielded similar results. A second improvement was achieved by coating the slides with 7% celloidin solution in ethyl alcohol-ether prior to AgNOR staining and acid pretreatment. This coating prevented nonspecific silver deposition on argyrophilic bacteria and other tissue debris in human vaginal smears that could make visualizing AgNOR sites difficult. Finally, placing sections face down on the staining solution prevents the formation of nonspecific silver precipitates. These procedures can be applied together or separately according to the requirements of the material to be evaluated.

Cellular Proliferation in Tumors: A Review of Methods, Interpretation, and Clinical Applications

Cellular proliferation with altered control is one of the 1st characteristics of a neoplastic cell population. Although tumor growth reflects both tumor cell replication and cell loss (opposing growth and antigrowth factors), most studies published in veterinary literature used immunohistochemistry (proliferating cell nuclear antigen [PCNA]; Ki-67 [MIB-1]) or staining for argyrophilic nucleolar organizer regions (AgNORs) to measure proliferation. These studies have appeared in the veterinary literature for more than a decade, describing associations between proliferation indices and histologic grade, biological behavior, and clinical outcome for some tumor types but no clinically relevant associations for other tumors. The results of these studies are summarized here. Methods for evaluation of the numerous regulatory proteins that direct the cell throughout its cycle are now available and should allow more precise identification of cellular aberrations in cancer.

Long Term Pulmonary Toxicity of Indium Arsenide and Indium Phosphide Instilled Intratracheally in Hamsters

Long Term Pulmonary Toxicity of Indium Arsenide and Indium Phosphide Instilled Intratracheally in Hamsters: Koji Yamazaki, et al. Department of Hygiene, Graduate School of Medical Sciences, Kyushu University—We examined the long‐term toxicological effects of III‐V semiconductor particles on laboratory animals. Eight‐week‐old male Syrian golden hamsters were given 4 mg/kg indium arsenide (InAs) or 3 mg/kg indium phosphide (InP) particles, both containing 2.4 mg/kg as indium, intratracheally twice a week for 8 weeks. Control hamsters were given only a vehicle, phosphate buffer solution. Over a 2‐yr period, these animals were euthanized serially and the biological effects were determined. Weight gain was significantly suppressed in both InAs and InP groups, compared to the control group, with greater suppression in the InAs group. The serum indium concentration in the InAs group was about twice as high as that in the InP group, in each period. Histopathologically, severe pulmonary inflammation and localized lesions with bronchiolo‐alveolar cell hyperplasia were present in both InAs and InP groups from just after the last administration. The localized lesions gradually transformed to proteinosis‐like lesions with periodic acid Schiff reagent positive exudation after 16 wk. By means of immunostaining of proliferating cell nuclear antigen and argyrophilic proteins associated with nucleolar organizer regions staining, proliferative activities were evidenced in the localized lesions at each time and were noticeable in their early stage. K‐ras, a known oncogene, was not mutated in association with these lesions. In conclusion, InAs and InP particles caused severe systemic toxicity and pulmonary localized hyperplastic lesions with proliferative activity were derived via the respiratory route. Neoplastic change was nil even in a 2‐yr observation period.

Morphological Changes in Silver-stained Nucleolar Organizar Regions (AgNORs) Related to Proliferative Activity in Glutathione S-transferase Placental From (GST-P) Positive Foci of Rat Liver.

Silver (Ag)-staining nucleolar organizer region (NOR) proteins are closely related to the cell cycle and reflect proliferative activity. In this study, we examined morphological changes of AgNORs in both glutathione S-transferase placental form (GST-P) positive foci and background parenchyma of rat liver by double staining. To induce GST-P positive foci, eight male F344 rats were given a single injection of diethylnitrosamine (20 mg/kg b.w., i.p.), and fed a diet containing 0.01% 2-acetylaminofluorene in diet from weeks 2 to 4 and subjected to 2/3 partial hepatectomy at week 3. At the end of week 5, liver sections were processed for double staining of GST-P and AgNOR. No statistically significant difference was observed for the number of AgNORs between GST-P positive foci and background parenchyma. However, areas of AgNORs relative to nuclear area (dot-to-nuclei ratio) in GST-P positive foci were significantly increased and the roundness was decreased as compared to values for surrounding hepatocytes. Frequencies of cells with big and bizarre (low roundness) dots were elevated in GST-P positive foci. Hence, our data suggest that AgNOR area and shape rather than number may reflect cell proliferation more acurately, so that assessment of these parameters may be useful for evaluating the biological nature of preneoplastic lesions.

Study of borderline and invasive mucinous ovarian tumors using Ki-67 (MIB 1) antibodies and nucleolar organizer region (NOR) staining.

Extensive application of new methods in histopathology has resulted in large quantities of data on molecular markers in different types of human tumors. The main purpose has been to obtain additional parameters for the characterization of various types of malignancy to give more precise information on their biological behavior. In this study we tested AgNOR and Ki-67 (MIB 1) staining in 39 ovarian mucinous tumors to evaluate its diagnostic potential in distinguishing between borderline tumors and well-differentiated carcinomas at stage I of FIGO classification. In general, there was an increase in AgNOR median values from benign (1.83 +/- 0.8) to well-differentiated carcinomas (2.91 +/- 1.3) with an intermediate value in borderline (2.3 +/- 1.6). The lowest Ki-67 labeling index (LI) was found in benign cystadenomas (5.7% +/- 3.4%) and papillary mucinous cystadenomas (7.9 +/- 5.8%). In borderline tumors mean LI was 12.2 +/- 10.9% whereas well-differentiated carcinomas had higher LI (16.8 +/- 10.2%). There were no statistically significant differences between the evaluated tumors and techniques applied. In our opinion the AgNOR count and Ki-67 LI do not appear useful for assessing differences between borderline and well-differentiated mucinous ovarian tumors at stage I of clinical advancement.

Potentially malignant and malignant lesions of the lip. Role of silver staining nucleolar organizer regions, proliferating cell nuclear antigen, p53, and c-myc in differentiation and prognosis.

The cellular changes leading to carcinoma of the lip are still not completely understood. This study was carried out on 44 malignant and potentially malignant lesions of the lower lip [30 squamous cell carcinomas (SCC), 7 actinic cheilitis, 3 leukoplakias, and 4 nodal metastases from lower lip SCC]. Silver-stained nucleolar organizer regions (AgNORs) and the immunohistochemical expression of proliferating cell nuclear antigen (PCNA), p53, and c-myc were evaluated on formalin-fixed, paraffin-embedded sections. The results indicate that the size and numbers of AgNORs and the percentage of PCNA-positive cells are sensitive parameters for discriminating between potentially malignant lesions and SCC, and for the prognostic sub-typing of lower lip SCC. Furthermore, while p53 positivity was found more frequently in high-grade carcinomas, p53-positive cellular clones were also found in some potentially malignant lesions, a finding probably related to ultraviolet-related cellular damage. These p53-positive lesions could be considered at higher risk of progression to malignancy than the p53-negative ones, although there is no evidence for this as yet. c-myc positivity was found only in some high-grade carcinomas and metastases, and appeared correlated with the later phases of lip carcinogenesis. The combined evaluation of the proliferation status, together with the changes in p53 and c-myc oncoproteins, might constitute useful markers for the prognostic evaluation of potentially malignant, as well as malignant, lesions of the lip.

Cytotaxonomic studies of Encarsia Förster (Hymenoptera: Aphelinidae)

AbstractA cytotaxonomic study was carried out on 13 species of Encarsia Förster, known to parasitize aleyrodids and diaspidids. The chromosomes varied greatly both in number and morphology, with E. protransvena Viggiani having the lowest chromosome number (2n = 6) and E. asterobemisiae Viggiani & Mazzone the highest (2n = 20). The most common chromosome number was 2n = 10. C-banding, G-banding and silver staining for nucleolar organizer regions (NOR) provided the possibility of distinguishing between karyotypes with the same chromosome number and morphology and to identify all the pairs of homologues in a diploid set for advanced cytogenetic studies. The karyotype of Coccophagus lycimnia (Walker) was also examined. The chromosome data suggested that in Encarsia, karyotype differentiation has mainly occurred through a series of centric fusions, although other rearrangements may also have been significant. The importance of karyological data in systematic studies and in the identification of biotypes and cryptic species of economic interest is outlined.

Nucleolar organizer regions (Ag-NORs) in multinuclear osteoclasts

Osteoclasts are multinuclear bone-resorbing cells. Little is known about regulation of gene expression in these cells and how it changes during the aging of an osteoclast, which is indicated by an increasing number of nuclei. Silver staining of nucleolar organizer regions, so-called Ag-NORs, allows us to assess the activity of rRNA expression. Using this technique, we show that individual nuclei of osteoclasts can be stained specifically and that the majority of individual nuclei contained Ag-NORs. The number and size of Ag-NORs varied, indicative of difference in the progression of the cell cycle through the G1 phase. The relative activity of Ag-NORs was not synchronized on the level of individual nuclei, regardless of the number of nuclei per osteoclast. Surprisingly, the total activity of Ag-NOR appeared to be correlated to the total nuclear area of an osteoclast. We conclude that regulation of gene expression in multinuclear osteoclasts most likely occurs on the level of individual nuclei.

Heat shock protein expression and drug resistance in breast cancer patients treated with induction chemotherapy.

Heat shock proteins (Hsps) are induced in vitro by several cytotoxic drugs; in human breast cancer cells these proteins appear to be involved in anti-cancer drug resistance. The present report was designed to analyze whether chemotherapy affects in vivo the expression of Hsp27, Hsp70, Hsc70 and Hsp90 in breast cancer patients treated with induction chemotherapy and whether these proteins may be determinants of tumor resistance to drug administration. We have analyzed 35 biopsies from breast cancer patients treated with induction chemotherapy. Expression of the Hsps in the tumors was compared with (i) histological and clinical responses to chemotherapy, (ii) tumor cell proliferation measured by proliferating cell nuclear antigen (PCNA) immunostaining and nucleolar organizer regions (AgNORs) staining and (iii) the expression of estrogen and progesterone receptors. We also compared disease-free survival (DFS) and overall survival (OS) with the expression of the Hsps studied. After chemotherapy, nuclear Hsp27 and Hsp70 expression was increased and Hsp70 and Hsc70 cytoplasmic expression was decreased. A high nuclear proportion of Hsp70 in tumor cells (>10%) correlated significantly with drug resistance. We also observed that patients whose tumors expressed nuclear or a high cytoplasmic proportion (>66%) of Hsp27 had shorter DFS. The combination of Hsp27 and Hsp70 levels showed a strong correlation with DFS. Neither the cellular proliferation nor the levels of steroid receptors showed any significant difference before or after drug administration or during follow-up of patients. Our results suggest that Hsp27 and Hsp70 are involved in drug resistance in breast cancer patients treated with combination chemotherapies.

Heat shock protein expression and drug resistance in breast cancer patients treated with induction chemotherapy

Heat shock proteins (Hsps) are induced in vitro by several cytotoxic drugs; in human breast cancer cells these proteins appear to be involved in anti-cancer drug resistance. The present report was designed to analyze whether chemotherapy affects in vivo the expression of Hsp27, Hsp70, Hsc70 and Hsp90 in breast cancer patients treated with induction chemotherapy and whether these proteins may be determinants of tumor resistance to drug administration. We have analyzed 35 biopsies from breast cancer patients treated with induction chemotherapy. Expression of the Hsps in the tumors was compared with (i) histological and clinical responses to chemotherapy, (ii) tumor cell proliferation measured by proliferating cell nuclear antigen (PCNA) immunostaining and nucleolar organizer regions (AgNORs) staining and (iii) the expression of estrogen and progesterone receptors. We also compared disease-free survival (DFS) and overall survival (OS) with the expression of the Hsps studied. After chemotherapy, nuclear Hsp27 and Hsp70 expression was increased and Hsp70 and Hsc70 cytoplasmic expression was decreased. A high nuclear proportion of Hsp70 in tumor cells (>10%) correlated significantly with drug resistance. We also observed that patients whose tumors expressed nuclear or a high cytoplasmic proportion (>66%) of Hsp27 had shorter DFS.<0B> <0R>The combination of Hsp27 and Hsp70 levels showed a strong correlation with DFS. Neither the cellular proliferation nor the levels of steroid receptors showed any significant difference before or after drug administration or during follow-up of patients. Our results suggest that Hsp27 and Hsp70 are involved in drug resistance in breast cancer patients treated with combination chemotherapies. Int. J. Cancer (Pred. Oncol.) 79:468–475, 1998.© 1998 Wiley-Liss, Inc.

Possible Prognostic Markers in Conjunctival Dysplasia and Squamous Cell Carcinoma

Conjunctival squamous cell carcinoma (SCC) can develop from carinoma in situ or severe dysplasia known as conjunctival intraepithelial neoplasia (CIN). Conjunctival intraepithelial neoplasia and SCC are histopathologically well-defined conditions. However, it is difficult to determine the grading of dysplasia by clinical morphologic findings. Recently, proliferating cell nuclear antigen (PCNA) immunostaining, p53 immunostaining, and argyrophilic nucleolar organizer regions (AgNORs) staining have been established as valuable means of studying the biologic behavior of malignant cells. In the present study, these three staining techniques were used to examine histologic preparations of three conjunctival dysplasia and one SCC lesion. Five conjunctival tumor samples were obtained from four patients between July 1993 and October 1995. Following formalin fixation and embedding in paraffin, PCNA, and p53 immunostaining and AgNORs staining was performed with all tissue specimens. The PCNA-positive rate was the highest in SCC, followed by severe dysplasia and mild dysplasia. The p53-positive rate was the highest in severe dysplasia, followed by mild dysplasia, and negative in SCC. The AgNORs-count increased as malignancy advanced. These staining methods, which are markers for proliferative potency and cell differentiation, will be useful for early detection of changes in malignancy and will aid in decisions on treatment and prognosis.

A study of the proliferative activity of the long junctional epithelium using argyrophilic nucleolar organizer region (AgNORs) staining.

The proliferative activity of the long junctional epithelium (LJE) in rats was examined using stains for argyrophilic proteins of the nucleolar organizer region (AgNORs protein). The LJE was experimentally produced by insertion of a rubber piece between maxillary molars for 1 wk. After removal of the rubber, the length and AgNORs parameters of the LJE were measured and analyzed statistically. The LJE widely covered the apical side of the exposed root surface 4 wk after the removal. Its length was longest after 4 and 8 wk; it became shorter subsequently. The AgNORs were visible as black dots of various sizes and numbers on the sections. A high potential for proliferation was obvious in the LJE after 4 wk and was maintained until 12 wk after the removal. The AgNORs ratio on the connective tissue interface of the LJE was about twice of that of normal junctional epithelium after 4-12 wk. The proliferative activity on the root surface side was slightly increased after 4 wk. There was no significant difference in proliferative activity between the coronal and apical sides. These results suggest that the proliferative activity of the LJE is maintained continuously at a high level on the connective tissue interface supplying the epithelial cells. Basal cells proliferate at the connective tissue interface of the LJE, migrate directly to the root surface or via the apical portion and finally desquamate from the surface of the epithelium.

A Comparison of Triploid Induction Validation Techniques

Abstract Triploidy induction is a technique that allows genetic manipulation of chromosome number to control reproduction and potentially create faster-growing animals; however, most methods for inducing polyploidy are not 100% effective. Using sunshine bass (white bass Morone chrysops ♀ × striped bass M. saxatilis ♂) as a model, we cross-validated the most common verification techniques: DNA staining and fluorescence quantification with a flow cytometer, erythrocyte nuclear volume with a Coulter counter particle size analyzer, silver staining of nucleolar organizer regions (NORs), and cytological karyotyping. Results indicated that the electronic techniques of particle size analysis and flow cytometry were the simplest and quickest methods of validation. The major drawback of both electronic ploidy determination methods is the cost of the equipment required for analysis. Cytological karyotyping was the most accurate method for determining polyploidy because actual chromosome numbers were determined. It was also the most time-consuming, tedious, and frustrating of the techniques, which reduces its applicability in mass screening of fish. Silver staining was the least expensive technique used for verifying a nominal number of fish, but it was also the most suspect because the NORs were sometimes difficult to detect, and there were conflicting results in older fish. All techniques demand a certain technical competence that can either be self-taught or requires extramural training.

A study of the proliferative activity of the long junctional epithelium using argyrophilic nucleolar organizer region (AgNORs) staining

The proliferative activity of the long junctional epithelium (LJE) in rats was examined using stains for argyrophilic proteins of the nucleolar organizer region (AgNORs protein). The LJE was experimentally produced by insertion of a rubber piece between maxillary molars for 1 wk. After removal of the rubber, the length and AgNORs parameters of the LJE were measured and analyzed statistically. The LJE widely covered the apical side of the exposed root surface 4 wk after the removal. Its length was longest after 4 and 8 wk; it became shorter subsequently. The AgNORs were visible as black dots of various sizes and numbers on the sections. A high potential for proliferation was obvious in the LJE after 4 wk and was maintained until 12 wk after the removal. The AgNORs ratio on the connective tissue interface of the LJE was about twice of that of normal junctional epithelium after 4‐12 wk. The proliferative activity on the root surface side was slightly increased after 4 wk. There was no significant difference in proliferative activity between the coronal and apical sides. These results suggest that the proliferative activity of the LJE is maintained continuously at a high level on the connective tissue interface supplying the epithelial cells. Basal cells proliferate at the connective tissue interface of the LJE, migrate directly to the root surface or via the apical portion and finally desquamate from the surface of the epithelium.

Effects of dexamethasone on pulp cells in cultured human teeth.

This study set out to examine in vitro the action of dexamethasone (Dex), a synthetic glucocorticoid on the pulp cells of human teeth, particularly in the cell-rich zone of the pulp where cell density is especially high. The present experiment was performed on 10 third molars extracted from patients aged between 16 to 20 years for orthodontic reasons under local anesthesia. The teeth were otherwise found to be sound and free of dental caries. Upon extraction, each tooth was divided into two parts mesio-distally. Each part was then cultured separately in Dulbecco's modified Eagle medium supplemented with 10% fetal bovine serum for a period of 7 days. 100nM Dex, 1mM sodium-β-glycerophosphate and 0.2mM ascorbic acid were added to one half of the total cultures which served as the experimental group. Four areas of the pulp tissue, the roof, the root, the lateral wall and the base were then each divided into three layers of equal thickness measuring 150×33.0±8.26 (mean±S.D.) μm to correspond to the average thickness of the cell-rich zone and observed histological examinations of the cells were performed by hematoxylin-eosin (H.E.) staining and silver-binding nucleolar organizer region (AgNOR) staining. Results obtained from the test group cells showed a significant increase in the mean number of AgNOR in the roof and root areas in the cell-rich zone in addition to a slightly larger mean cell size in almost every zone. This shows that culture of 7 days in dexamethasone-added medium can lead to a heightened level of cell activity in the roof and root areas of the cell-rich zone.

The gastric hypercellular microleiomyoma as a precursor lesion for clinical gastrointestinal stromal tumors

Differentiation features and proliferation activity of 67 gastric microleiomyomas (microLMs) and 53 clinical gastrointestinal stromal tumors (GISTs) of the stomach were compared. The 67 microLMs were divided into two categories on the basis of cellularity: 53 hypocellular and 14 hypercellular types, and the 39 GISTs were divided into 13 low-grade and 40 high-grade lesions. Immunohistochemically, 49 hypocellular microLMs (92%) were positive for alpha-smooth muscle actin and desmin, whereas only 16 (30%) were stained for vimentin. Conversely, all 14 hypercellular microLMs were positive for vimentin, and only one (7%) was positive for alpha-smooth muscle actin and desmin. Five low-grade (38%) and 14 high-grade GISTs (35%) were positive for alpha-smooth muscle actin, and 12 low-grade (92%) and all 40 high-grade GISTs were stained for vimentin. CD34 was positive in 10 hypocellular microLMs (19%), all 14 hypercellular microLMs, 10 low-grade GISTs (77%), and 38 high-grade GISTs (95%). Hypercellular microLM thus showed similarities to clinical GIST and also exhibited significantly higher proliferation activity than hypocellular microLM on analysis of the Ki-67 labeling index and argyrophilic nucleolar organizer regions staining. The findings indicate that the hypercellular microLM may be a direct precursor for clinical GIST, both showing a primitive mesenchymal cell nature.

Diagnostic importance of the nucleolar organizer regions in Wegener's granulomatosis.

The number and distribution pattern of silver staining nucleolar organizer regions (AgNORs) were thought to reflect the cellular proliferative activity and the malignancy of tumours. Using a silver-staining method, the variations of AgNORs have been studied in patients with atypical inflammatory lesion (n = 5), malignant reticulosis (n = 5) and Wegener's granulomatosis (n = 6). Our results reveal that there was a statistically significant difference (p < 0.01), highly suggestive of a difference in AgNOR counts between the atypical inflammatory lesion and Wegener's granulomatosis, with the Wegener's granulomatosis specimens having the higher irregular AgNORs, but the difference between Wegener's granulomatosis and malignant reticulosis is probably not clinically important. It is concluded that AgNORs may be useful in differentiating Wegener's granulomatosis from an atypical inflammatory lesion, and the simplified counting technique is adequate for the purpose.

Usefulness of Argyrophilic Nucleolar Organizer Regions Score to Differentiate Suspicious Malignancy in Pulmonary Cytology

Pulmonary cytologic specimens reported as "suspicious for malignancy" pose problems in clinical management. Silver staining for argyrophilic nucleolar organizer regions (AgNOR) has proved useful in making a cytopathologically differential diagnosis between benign and malignant cells. This study aimed to evaluate the usefulness of AgNOR score in the diagnosis of pulmonary cytologic specimens deemed inconclusive by conventional staining methods. Pulmonary cytologic specimens initially reported as suspicious for malignancy with Papanicolaou or May-Grünwald-Giemsa (MGG) staining obtained from 35 proved cases were destained then restained using the AgNOR technique. Another 35 cases with clear cytologic diagnosis were also examined for comparison. The median number of dots, defined as the AgNOR score, was used to differentiate malignant from benign specimens. Malignant cases had significantly higher AgNOR scores than benign ones (p<0.001). There were no significant differences among smears previously stained with Papanicolaou or MGG method, among specimens obtained via bronchoscopic brushing, fine-needle aspiration of lung or pleural effusion, or among subgroups of malignant diseases. Based on the results of our previous study, the cutoff value of the AgNOR score to differentiate benignancy from malignancy was set at 6. At this setting, the sensitivity and specificity of AgNOR score were 88% and 80%, respectively, in aiding a differential diagnosis of pulmonary cytologic specimens initially classified as suspicious for malignancy. For those cases with a clear cytologic diagnosis, the sensitivity and specificity of AgNOR score were 92% and 100%, respectively. For all cases, the sensitivity of AgNOR score was 90% and the specificity was also 90%. The AgNOR score is of value in aiding a differential diagnosis between benign and malignant lesions in pulmonary specimens with equivocal cytologic features.

Usefulness of AgNOR score in differentiating benign from malignant pulmonary aspiration cytology.

Malignant cells are known to display greater argyrophil staining for nucleolar organizer regions (AgNORs) than for benign cells due to their active proliferation. In this study we assessed the diagnostic value of AgNOR staining on 47 fine needle aspiration cytologic specimens of lung previously stained with the May-Grünwald-Giemsa (MGG) method. Cytologic specimens obtained from fine needle aspiration of the lung in 47 proven cases prestained with the MGG technique were destained and restained with the AgNOR method. Seventeen of them were benign and 30 malignant. To differentiate malignant from benign entities, the highest median value for AgNOR number (AgNOR score) obtained from the benign cases was chosen as a cutoff point (test specificity, 100%). AgNOR scores of malignant cases were significantly higher than those of benign cases (P < .001). There was no significant difference between two subgroups of benign diseases or among four subgroups of malignant diseases. The sensitivity of the AgNOR score was 93% (28/30) in providing a diagnosis of malignancy when the cutoff value was set at 6. The AgNOR technique may be of considerable value in aiding a diagnosis of malignancy, especially when the score is > 6.

Radiation and speciation of spider monkeys, genus Ateles, from the cytogenetic viewpoint

The chromosomes of 22 animals of four subspecies of the genus Ateles (A. paniscus paniscus, A. p. chamek, A. belzebuth hybridus, and A. b. marginatus) were compared using G/C banding and NOR (nucleolar organizer region) staining methods. The cytogenetic data of Ateles in the literature were also used to clarify the phylogenetic relationships of the species and subspecies and to infer the routes of radiation and speciation of these taxa. Chromosomes 6 and 7 that showed more informative geographic variation and the apomorphic form 4/12, exclusively in A. p. paniscus, are the keys for understanding the evolution, radiation, and specification of the Ateles taxa. The ancestral populations of the genus originated in the southwestern Amazon Basin (the occurrence area of A. paniscus chamek) and spread in the Amazon Basin and westward, crossing the Andes and colonizing Central America and northwesternmost regions of South America. The evolutionary history of the northern South American taxa is interpreted using the model of biogeographical evolution postulated by Haffer [Science 185:131–137, 1969]. Ateles paniscus paniscus is the genetically most differentiated form and probably derives from A. belzebuth hybridus. Based on the karyotype differences, the populations of Ateles can be divided into four different group. These findings indicate the necessity of a more coherent taxonomic arrangement for the taxa of Ateles. Am. J. Primatol. 42:167–178, 1997. © 1997 Wiley-Liss, Inc.