Karyo-Morphology and Nucleoli Analysis of Commelina L. (Commelinaceae) from Ethiopia

Karyo-Morphology and Nucleoli Analysis of Commelina L. 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(Commelinaceae) from Ethiopia Samuel Gebrechristos Gebretsadkan, Gebrehiwet Kiros, Bahlibi Gebreabzgi, and 2 more This is a preprint; it has not been peer reviewed by a journal. https://doi.org/ 10.21203/rs.3.rs-100645/v1 This work is licensed under a CC BY 4.0 License Status: Posted Version 1 posted You are reading this latest preprint version Abstract Background With about 100 species, Commelina is the largest genus of Commelinaceae in Africa. Although medicinal and economic benefits had been studied extensively, little is known about its cytological analysis. Hence, this study will focus on chromosome and nucleoli analysis of selected species of Commelina L. Somatic chromosomes were prepared from root tips that emerged from the nodes of stem cuttings that were made to stand submerged in water. The roots were pretreated in 8-hydroxyquinelin 3-5 hrs followed by fixation in 3:1 ethanol: acetic acid for 1-24 hrs at 4 O C. Air-dry slides were prepared following cellulase and pectinase maceration at 37 O C, the preparation was stained in Giemsa stain (PH 6.4), rinsed and mounted. Nucleoli were stained in silver nitrate solution. Results Chromosome numbers and Karyotype formula of the four species were found as C. africana 2 n =2 x =30 (12m + 10sm + 8st), C. benghalensis 2 n =6 x =66 (36m + 24sm + 6st), C. diffusa (Ginchi) 2 n =66 (28m +26sm + 12st), C. diffusa ( Jimma ) 2 n =2 x =30 (10m + 8sm + 12st) and C. subulata 2 n =2 x =30 (18m +10sm + 2st). According to Stebbins karyotype asymmetry, the karyotypes of C. africana and C. subulata were 2A type, while that of C. benghalensis and C. diffusa (Ginchi) were 2B type. 3A asymmetry type was obtained for C. diffusa (Jimma). Karotypes of Tradescantia were found to be monomodal for the Commelina species of the studied plant materials. Satellites were observed for species C. africana and C. diffusa with variation in number ranging from 2 to 6. The maximum number of nucleoli observed varies from two to four for Commelina . Conclusions  This study reported karyotype and nucleoli of the Ethiopian Commelinaceae for the first time. The current investigation can be considered as an additional karyotype data to the earlier meiosis report for Ethiopian materials. Molecular Genetics Commelina L. Ethiopia Karyotype Nucleoli Satellite Figures Figure 1 Figure 1 Figure 2 Figure 2 Background The family Commelinaceae, commonly known as the spiderwort, is known to possess 41 genera and 650 species throughout the world [ 1 ], with main distribution in tropics and subtropics that extends into northern temperate regions. The family is represented by 9 genera and 56 species in Ethiopia [ 2 ].With about 100 species, Commelina is the largest genus of Commelinaceae in Africa. At least 65 species occur in the combined areas of the Flora of Tropical East Africa (Kenya, Uganda, and Tanzania) and Flora Zambesiaca (Malawi, Mozambique, Zambia, Zimbabwe, and Botswana) [ 3 , 4 ]. About 19 species have been identified in Ethiopia and Eritrea including 2 un-described species that are endemic to Ethiopia [ 5 ]. The genus Commelina has several medicinal significances. C. benghalensis is used to treat bed sores, breast sores and pimples in Pakistan [ 6 ]. In East Africa, the sap of C. benghalensis leaves and stems is used to treat ophthalmia, infertility in women, leprosy, sore throat and burns, and the liquid contained in the flowering spathe is used to treat eye complaints in Zanzibar [ 7 ]. People from Nepal use a paste derived from the plant to treat burns, and indigestion with a juice produced from the roots [ 8 ]. In China, C. diffusa is used as a medicinal herb with febrifugal and diuretic effects [ 9 ]. C. benghalensis is known for its medicinal purposes as they have already been reported in treating hypertension, burns, leprosy, sore throats, cataplasm, and wound healing [ 10 ]. Numerous compounds have been identified from the vegetative and flower parts of Commelina benghalensis including noctacosanol, n-triocotanol, stigma-sterol, compesterol and hydrocyanic acid [ 11 ]. Phyto-chemical screening also revealed the presence of many secondary metabolites like phlobatannins, carbohydrates, tannins, glycosides, volatile oils, resins, balsams, flavonoids and saponins [ 12 ]. Presence of flavonoids, for example, indicates the plant might have an antioxidant, anti-allergic, anti-inflammatory, anti- microbial or anti- cancer activity [ 13 ]. In addition, C. benghalensis has analgestic action that proves the folkloric use in pain management [ 14 ]. Although several medicinal and economic uses of the family Commelinaceae are known, little has been done in cytological analysis in Ethiopia. Previous research done only focused on chromosome number of four species included in this study based on meiosis and voucher specimen were taken only from the Harar Province except for Cyanotis barbata D. Don which was from Debrezeit [ 15 ]. The present study was focused on chromosome number, karyotype analysis and silver staining. Here the chromosome number, ploidy level, karyotype and number of nucleoli organizing region (NORs) of these species of Commelinaceae are described. Such study can help to expand the current cytological knowledge of the Ethiopian Commelinaceae with further contribution for phylogenetic and biosystematics research. Results The Commelina species contain metacentric, sub-metacentric and sub-telocentric chromosomes with variation in number and morphology (Table 1). C. africana Somatic chromosomes observed from more than five intact cells showed that C. africana collected from Addis Ababa contained a diploid number of 2 n =2 x = 30. Because of its better chromosome spread and clear chromosome morphology a cell with 29 chromosome number is shown in Figure 1A and a karyotype constructed from same cell in Figure 1B. This variant of the species consisted of six pairs of metacentric chromosomes (pairs 1 to 6), five pairs of sub- metacentic chromosomes (pair numbers 7 to 11) and four sub-telocentrics (pairs 12 to 15) (Figure 1B). The karyotype formula is, therefore, 12m + 10sm + 8 st (Table 1). According to Stebbins karyotype classification, this species belongs to 2A type with asymmetry index of 65.19 (Table 1). Small satellites were also observed at the tip of the short arm of a pair of chromosomes (Figure 2A) and these correspond to the 8 th pair on the karyotype (Figure 1B). Figure 2B shows three interphase nucleoli, from which it can be assumed that the maximum number of nucleoli and so chromosome satellites for this species is four. Length wise measurement of metaphase chromosomes revealed a value range from the smallest 4.53µm to largest 7.5µm (Table 1). Total length of whole chromosome complement (2 n ) was 176.72µm. The chromosomes were medium in size with an average length and asymmetry of 5.89µm and 1.87, respectively. C. benghalensis According to somatic chromosome analysis of the root tips, this species possesses 2 n =6 x =66 (Figure 1C and Table 1). The karyotpe constructed based on calculated arm ratio and chromosome size revealed the presence of eighteen metacentric chromosome pairs (pair numbers 1, 3, 4, 5, 8, 11, 14, 15, 16, 18, 22, 24, 25, 29, 30, 31, 32, 33), twelve sub-metacentrics (pair numbers 6, 7, 9, 10, 12, 17, 19, 20, 21, 23, 27, 28) and 3 sub-telocentric pairs (pairs 2, 13, 26 ) (Figure 1D). The karyotype formula was, therefore, 36m + 24sm + 6st. As to Stebbins method of classification, C. benghalensis possesses asymmetry index of 62.58 with 2B in karyotype asymmetry (Table 1). A maximum of five nucleoli were observed through silver staining but no satellites were observed (Figure 1E). Two of the nucleoli are much larger than the other three nucleoli. Each of the larger nucleoli might have resulted from fusion of two or more nucleoli. It can be inferred that six or more chromosome satellites are present in this species. The chromosome lengths differ from the smallest 2.5µm and gradually increased up to the largest 5.625µm (Table 1). The total length of whole chromosome complement was about 267.935µm (Table 1). The total length of the long arms of the diploid set was 167.675µm and that of short arms for diploid set was 100.26µm with their ratio generating an average karyotype asymmetry of 1.672. Generally, C. benghalensis (Addis Ababa) has predominantly metacentric and sub-metacentric type of chromosomes. The chromosomes were short to medium in size with an average length of 4.06µm. C. diffusa from Ginchi The variant of C. diffusa collected from Ginchi possessed 2 n = 66 (Table 1 and Figure 1E). Based on centromeric ratio and chromosome length measurements , the karyotype (Figure 1F) consisted of fourteen metacentric pairs (pair numbers 1, 4, 6, 15, 16, 20, 21, 23, 25, 26, 30, 31, 32, 33), thirteen sub-metacentrics (pairs 2, 3, 5, 7, 8, 9, 10, 13, 17, 19, 24, 27, 28) and six sub-telocentrics (pairs 11, 12, 14, 18, 22, 29) with a formula of 28m + 26sm + 12st. The asymmetry index is 65.56 and belongs to 2B type of Stebbins karyotype classification (Table 1). As shown in Figure 2(C and D), a total of six satellites and four nucleoli were observed from chromosome preparation and silver staining, respectively. Size of satellites varies with one pair being larger than the other two. C. diffusa (Ginchi) has comparatively larger chromosomes than the other Commelina species included in this study with a length range from smallest 4.375µm to largest 11.25µm and total length of whole chromosome complement 507.53µm (Table 1). The total length of long arms and short arms for the diploid set was 332.745µm and 174.785µm respectively (Table 1). The average karyotype asymmetry is 1.904 and on the average the chromosomes can be classified under sub-metacentric. The chromosomes were medium to large in size with an average length of 7.69µm. C. diffusa (Ginchi) has intra-chromosomal and inter-chromosomal asymmetry indices of 0.435 and 0.17035 respectively (Table 1). This variant also has a coefficient of variation in chromosome length of (17.035), ratio of centromeric gradient (53.0163) and dispersion index (9.031) (Table 1). Table 1 Cytological analysis of Commelina L. Species Chromosome length (µm) Shortest: longest ACL (µm) 2 n Basic number( x ) Ploidy level A 1 A 2 C v C G DI Karyotype Formula Number of metaphase plates Asymmetry index Stebbins asymmetry K L M C. africana 4.53 – 7.5 5.89 30 15 2x 0.4345 0.1498 14.98 55.32 8.29 12m + 10sm + 8st 10 65.19 0.43 1.67 2A C. benghalensis 2.5 – 5.625 4.06 66 11 6 x 0.3715 0.1582 15.82 57.831 0.3715 36m + 24sm + 6st 4 62.58 0.35 2.25 2B C.diffusa (Ginchi) 4.375 – 11.25 7.69 66 - - 0.435 0.17035 17.035 53.0163 9.031 28m + 26sm + 12st 5 65.56 0.5 2.57 2B C.diffusa (Jimma) 2.5 – 4.58 3.47 30 15 2 x 0.76717 0.13771 13.771 39.32 5.415 10m + 8sm + 12st 10 69.497 0.67 1.832 3A C. subulata 1.923 – 3.654 2.77 30 15 2 x 0.3523 0.1891 18.91 67.45 0.3523 18m + 10sm + 2st 4 60.232 0.267 1.9 2A ACL: average chromosome length; 2 n : somatic chromosome number; A1: intra-chromosomal asymmetry index; A2: inter-chromosomal asymmetry index; C v : Coefficient of variation of chromosome length; C G : ratio of centromeric gradient; DI: dispersion index K: percentage of chromosome with ratio=r>2; L: ratio of largest to smallest chromosome; M: degree of asymmetry C. diffusa from Jimma The material of this species collected from Jimma possessed 2 n =2 x = 30 (Table 1 and Figure 1G). According to centromeric ratio and chromosome length measurements, the karyotype (Figure 1H) has five metacentric pairs (pair numbers 1 to 5), 4 sub-metacentric pairs (pairs 6 to 9) and 6 sub-telocentric pairs (pair numbers 10 to 15) with a formula of 10m + 8sm + 12st. The asymmetry index is comparatively higher than the other studied Commelina species with a value of 69.497 and according to Stebbins karyotype asymmetry classification, C. diffusa (Jimma) belongs to 3A type (Table 1). Even if no satellites were observed from the current plant material, a total of three interphase nucleoli were observed that possibly predict the number of NORs or satellites to be four (Figure 2F). C. diffusa (Jimma) possesses the smallest chromosomes size next to C. subulata with a length range from shortest 2.5µm to longest 4.58µm and total length of whole 2 n chromosome complement 104.22µm (Table 1). The total length of long arms for the diploid set was 72.43µm and total length of short arms for the diploid set was 31.79. The chromosomes range from very small to medium in size with an average length of 3.47µm and karyotype asymmetry of 2.28µm. This particular variant has the highest value in terms of intra-chromosomal asymmetry index (0.76717). It also possesses the lowest inter-chromosomal asymmetry index (0.13771), coefficient of variation in chromosome length (13.771), ratio in centromeric gradient (39.32) and dispersion index (5.415) when compared to values from all other studied plants of the genus Commelina . C. subulata The experimental species collected from Ginchi indicated that C. subulata has a diploid number of 2 n =2 x = 30 (Table 1 and Figure 1I). As to arm ratio and chromosome length measurements, the karyotype (Figure 1J) consisted of nine metacentric pairs (pair numbers 1, 2, 3, 5, 8, 9, 10, 12, 13), five sub-metacentrics (4, 6, 11, 14, 15) and one pair of sub-telocentrics (7) with a karyotypic formula of 18m + 10sm + 2st. The species possesses the lowest value in asymmetry index (60.232), being the 2A type in Stebbins asymmetry classification (Table 1). Two interphase nucleoli were observed (Figure 2G), but no satellites were observed. One of the two nucleoli is larger than the other, which could result from fusion of two or more nucleoli; and it may be inferred that a total of four satellite chromosomes/ NORs are present. Commelina subulata (Ginchi) has the smallest chromosome size than other species included in this study with a length range from smallest1.923µm to largest 3.654µm and the total length of whole chromosome complement is 83.154µm (Table 1). . The total length of long arms and short arms for the diploid set was 50.085µm and 33.069µm, respectively (Table 1). The average karyotype asymmetry is 1.51 and the chromosomes, on the average arm ratio, can be considered as metacentric. The chromosomes are very small to medium in size with an average length of 2.77µm. C. subulata possesses the lowest intra-chromosomal asymmetry index, highest ratio of centromeric gradient and dispersion index with a value of 0.3523, 12.755 and 67.45, respectively. The species also consisted of an inter-chromosomal asymmetry index (0.1891) and coefficient of variation in chromosome length (18.91) (Table 1). Discussion In the four species studied from the genus commelina , base numbers of x =11 and x =15 have been obtained. Base numbers x =11, 13 and 15 was also previously reported for Commelina [ 16 , 17 ].The species Commelina benghalensis (2 n =6 x =66) was based on x =11 [ 18 ]. Others, C. africana , C. diffusa (Jimma) and C. subulata having chromosome number of (2 n =2 x =30) were based on x =15 base number. This result agrees with previous studies [ 15 , 17 , 19-24 ]. C. subulata of Ginchi has 2 n =30 and this was in agreement with other reports from materials collected from Ethiopia [ 15 ] and Ghana [ 25 ]. Tetraploid species of C. subulata 2 n =60 have also been reported for materials from India [ 26 , 27 ]. C. diffusa from Ginchi has 2 n =66 and this number is reported for the first time. Chromosome numbers of C. africana collected from Addis Ababa and Sebeta were diploids 2 n =30 and this was in agreement with previous reports [ 15 ]. Even if, the current count was diploid chromosome number, Morton [ 28 ] also found 2 n =28 for materials from Ghana and polyploids (2 n =60 and 2 n = 120) were also reported [ 15 ]. C. benghalensis of Addis Ababa was found to be hexaploids (2 n =6 x =66) and this was in agreement with earlier report [ 25 ]. Both diploid 2 n =22 and tetraploid (2 n =44) cytotypes have also been reported for C. benghalensis from materials of Nigeria, China, India, Japan, Uganda, Ethiopia and Tanganyika [ 15 , 18-21 , 23 , 25 , 29 , 30 ]. A diploid C. diffusa (2 n =30) has been found for specimens collected from Jimma. This result was in agreement with other reports [ 15 , 24 ]. Polyploid series of the West African C. benghalensis , C. africana and Aneilema umbrsum complexes are of autopolyploid origin based on observation of close similarity between polyploid and diploids with the absence of allied taxa which could have been involved in allopolyploidy [ 25 ]. A comparison made between the karyotypes of C. diffusa from Taiwan [ 23 ] and C. diffusa (Entoto mountain and Jimma) in the present study indicates that, although they share similar chromosome number (2 n =30), they differ in karyotypic detail. The karyotype formula of the Taiwan specimen was reported with only two chromosomal groups, m and sm, (10m + 20sm), while in the present materials three chromosomal groups (m, sm and st) with a formula of 16m+6sm+8st (Entoto) and 10m + 8sm + 12st (Jimma) were observed. Alam and Sharma reported variation in karyotypes among five populations of Indian C. diffusa having 2 n = 30 [ 21 ]. Four different karyotype formula within 2 n =30 chromosomes from India were also reported previously [ 20 ]. Factors, other than the chromosomal heteromorphism, like deviation in techniques of chromosome preparation, condensation difference and measurement technique can also lead to karyotype diversity among reports by different workers. These factors hinder comparison between various karyotypes and for real comparison mean values for each measurement of individual chromosomes must be taken. Furthermore, karyotype comparison for the remaining species was not performed as all the chromosome report matching to the current chromosome number had not been found. The chromosome report 2 n =28 for the two species of C. diffusa and C. africana was probably associated with the uncommon aneuploids formed due to loss of chromosome number [ 15 , 28 ]. But reduction in chromosome number can be associated with Robertsonian fusion [ 32 ] and translocation of all or most of its part followed by loss of the chromosome. High rate of prevalence of aneuploidy and polyploidy in the genus Commelina had reported [ 25 ]. Jones and Joplings reported the chromosome size of the genus Commelina as the smallest in the family [ 16 ]. Faden and Morton, on the other hand, confirmed the presence of medium to relatively large chromosome within the genus [ 17 , 25 ]. In the present study, except for C. subulata , the size was predominantly medium, whereas C. subulata has an average chromosome smaller than the medium size limit (3µm). The karyotype data of the present materials in the genus Commelina indicated the presence of three types of chromosomes with higher frequency of metacentric (m) and sub-metacentric (sm) types than sub-telocentrics (st). This was supported by earlier reports [ 17 , 25 ]. However the ratio of each chromosome type considerably varies between each species. A comparison made based on asymmetric index in the current study indicates that C. africana collected from Addis Ababa has same karyotype asymmetry with C. diffusa (Jimma) which possess 65.19% and 65.56%, respectively. C. benghalensis of the current study falls within 2B Stebbins category. This is in disagreement with the diploid material from Nigeria [ 30 ]. This might be associated with genetic difference that associated with difference in agro-ecology. The present study revealed some degree of variation in chromosome length between the studied species with no major chromosomal difference among themselves. This may be associated with differences in degree of chromosome condensation between the metaphase spread of different species measured. Thus, in practice, it is difficult to draw taxonomic conclusions simply by comparing chromosome length between taxa [ 33 ] unless one compares chromosomes condensed to same degree. Satellited chromosomes were observed frequently in Commelinaceae [ 25 ]. In the present study, satellites were observed in C. africana and C. diffusa (Ginchi). The reason why satellites were not observed in some of the species may be that the satellites are too small and escape easy cytological detection, or condense. All the satellites detected were located at the tip of the short arm of chromosome. Discrepancies in number of satellites reported for a particular taxon or population can be due to the inability to observe all the satellites because of variation in techniques of chromosome preparation, stages at the time of chromosome analysis and chromosomal polymorphism [ 34 ]. The maximum number of telophase nucleoli obtained through silver staining may correspond to the maximum number of satellites that are present in particular taxon [ 34 ]. Thus, even though all the satellites are not detected for various reasons discussed above, one can infer about the number of satellited chromosomes present in the taxon, if one is able to obtain the maximum number of nucleoli present. Accordingly it was assumed that the maximum number of satellites for C. africana will be four. Nucleoli are formed at NORs of the satellited chromosomes. During the cell cycle, nucleoli disappear at late prophase and reform during telophase. It is less easy to observe the maximum number of nucleoli during telophase. As the cell cycle proceeds from telophase to interphase, nucleoli tend to fuse together, and thus their number in most of the interphase nuclei is usually less than their number in telophase nuclei. Though it is less frequently, it is possible that the nucleus enters interphase without all the nucleoli being fused, in which case the maximum number of nucleoli can also be observed in interphase nuclei. The maximum number of nucleoli observed, be at telophase or interphase, can be used to infer about the number of active NORs the plant possesses. Even numbers of maximum nucleoli are expected because NORs chromosomes occur as homologous pair (s). In case the highest number observed is odd number, one may take the next higher even number as the number of nucleoli for the organism. Usually when odd number is observed, at least one of the nucleoli is larger than the rest of the nucleoli indicating that the large nucleolus is the product of fusion of smaller nucleoli. In the present study, maximum number of nucleoli observed for C. africana (Addis Ababa) is three, and for C. africana (sebeta) and C. diffusa (Entoto) is four each which allow to make an inference that they all have 4 NORs (satellite chromosomes). The number of nucleoli observed for C. benghalensis is five but two are very large relative to the other three nucleoli. The large ones are possibly fusion products of two or more small nucleoli. There are at least three pairs or more satellited chromosomes in this species. Conclusions The present study has revealed and confirmed chromosome number, ploidy level, karyotype and nucleolus numbers of six species of Commelinaceae which were collected from different localities of central and south western part of the country. Accordingly, this study showed that basic chromosome number for three species of Commelina ( C. diffusa, C. africana and C. subulata ) is x =15 and 2 n =30. The chromosomes of these species are predominantly of m and sm types. C. benghalensis (2 n =6 x =66) was also found to have x =11. The basic chromosome number for C. diffusa (Ginchi), 2 n =66 was different from previous reports and hence it could be another cytotype for the species. Variation in karyotype formula is also observed within species of C. africana and C. diffusa collected from different localities. C. diffusa (Ginchi) is vigour, longer with distinct morphology than the other diploid species. This is the first work to present karyotypes, satellite chromosomes and nucleoli of the Ethiopian Commelinaceae. Nevertheless an integrated data from karyotype, molecular study of chloroplast genome and evolution through considering more representative species is necessary to infer phylogenetic relationship among the taxa. Methods Four species of the genus Commelina which were collected from different localities of Central and Southwestern part of Ethiopia were analyzed cytologically. Chromosome study including numbers and morphology (karyotypes) was done on C-metaphase chromosomes of the root tip meristematic cells. Pretreatment To obtain clean roots for chromosome preparation, stems were harvested from potted plants and allowed to stand submerged in water. In a few days, roots emerged from the nodes of the submerged stems and harvested for metaphase arrest of mitosis. Hence, roots were immersed in 8-hydroxyquiniline (0.002M) for 3 to 5 hrs, fixed in 3:1 (v/v) of ethanol and glacial acetic acid 1-24 hr at about 4 o C. Maceration Roots from fixative were rinsed several times in distilled water and macerated in a solution of pectinase and cellulase in a water bath at 37 o C for about 1 hr or more. The process was stopped when the tips of the root started detaching from the root with or without agitation of the vial. Then, enzyme solution was carefully pipetted out and transferred to a watch glass or a petridish after rinsing. Air dry slide preparation One or more of the macerated root tips (depending upon the size) were pipetted on a clean slide and fresh fixative (3:1, ethanol: acetic acid) were added to the root tips, and the tips were mashed quickly with flat ended mounted needle. The slide was then allowed to air dry at room temperature and stored until needed for staining. Slide staining Slides with good preparations were screened under phase contrast microscope and good preparations were screened under phase contrast microscope before staining. The promising preparation was stained in Giemsa stain in Sorenson phosphate buffer solution (PH 6.8). When correct contrast of staining was obtained, the slides were rinsed in distilled water, and allowed to air-dry at room temperature at least for 24 hrs. The preparation was made permanent by mounting in depex mountant and the gum was allowed to set for several days following by examining under camera fitted microscope. Then, enlarged photomicrograph prints were made accordingly. Silver staining Slide preparation was similar with the above with the exclusion of treatment of 8-hydroxyquiniline. The latter treatment was omitted in order to obtain telophase cells during which the maximum number of nucleoli may be observed. Silver staining technique was similar to the work of Dagne and Heneen [ 35 ]. Karyotype analysis The printed pictures of chromosomes were scanned into computer and the lengths of the whole chromosomes and their arms were measured in terms of pixel per cm using micro measure computer software version 3.3. Accordingly, the arm ratio of the chromosomes was calculated by dividing the length of the long arm to that of short arm. Karyotypes were constructed by cutting and arranging the putative homologous chromosomes into pairs based on arm ratio( r ) and chromosome size using the Smart Type software version 0.8. Chromosomes were categorized into chromosome types based on arm ratio ( r ) according to [ 36 ] with slight modification. In the present case the term metacentric chromosome was used to include both M and m types with r =1.0-1.7. Sub-metacentric was used as synonymous with sub-median chromosome of r = 1.7-3.0 and st is similar to sub-telocentric when r =3.0-7.0. After accurate measurements of karyotyped chromosomes were obtained, intra-chromosomal asymmetry (A 1 ) and inter-chromosomal asymmetry (A 2 ) indices have been calculated [ 37 ]. Moreover, DI (dispersion index) of chromosomes was also calculated [ 38 ]. Measurements like intra chromosomal (A 1 ), inter-chromosomal (A 2 ), dispersion indices (DI) and Stebbins asymmetry depends on both chromosome size and centromeric position for estimating karyotype asymmetry of chromosomes. But asymmetry index depends only on centromeric position [ 39 ]. Declarations Ethics approval and consent to participate Not applicable Consent for publication Not applicable Availability of data and materials The data used for current study can be provided on request. Competing interests The authors declare that they have no competing interests Funding All chemicals and materials required for the current study were funded from an internal grant of Aksum University. Authors' contributions SG prepared the publication manuscript and all experimental works. GK, BG, GM and DG review all the work and incorporated necessary comments for better output. Acknowledgements It is our great pleasure to acknowledge Dr. Kifle Dagne and Professor Ensermu Kelbessa of Addis Ababa University, for their meticulous assistance. We would also like to extend our heartfelt appreciation to Addis Ababa University, School of Graduate Studies, Faculty of Life Sciences for permission to access laboratory facilities to carry out our research. References Kubitzki K, Rohwer J, Bittrich V: The families and genera of vascular plants, vol. 1: Springer; 1990. Kelbessa E, Demissew S: Diversity of vascular plant taxa of the flora of Ethiopia and Eritrea. Ethiopian Journal of Biological Sciences 2014, 13(Supp.)):37-45. Faden RB: New species of Commelina (Commelinaceae) from the Flora of Tropical East Africa. Novon 1994:224-235. Faden RB: New or misunderstood species of Commelina (Commelinaceae) from the Flora of Tropical East Africa and Flora Zambesiaca areas. Novon 2001:398-409. Edwards S, Demissew S, Hedberg I: Flora of Ethiopia and Eritrea: Hydrocharitaceae to Arecaceae: The National Herbarium, Addis Ababa University; 1997. Qureshi R, Bhatti GR: Ethnobotany of plants used by the Thari people of Nara Desert, Pakistan. Fitoterapia 2008, 79(6):468-473. Van der Burg W: Commelina benghalensis L. Plant resources of tropical Africa 2004, 2:212-214. Manandhar NP: Plants and people of Nepal: Timber press; 2002. Deyuan H, Robert A: Commelina diffusa. Wu, ZY; Raven, PH; Hong, DY, Flora of China 2000, 24. Fibrich B, Lall N: Commelina benghalensis. In: Underexplored Medicinal Plants from Sub-Saharan Africa. Elsevier; 2020: 77-85. Jayvir A: Nature Heals, A glossary of selected indigenous medicinal plants of India. Indian Journal of Pharmacology 1998, 30(2):126. Ibrahim J, Ajaegbu VC, Egharevba HO: Pharmacognostic and phytochemical analysis of Commelina benghalensis L. Ethnobotanical Leaflets 2010, 2010(5):7. 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FUKUMOTO K: EXAMINATION OF CHROMOSOME NUMBERS AND KARYOTYPES OF SOME COMMELINA SPECIES AND FORMAE. In: JAPANESE JOURNAL OF GENETICS: 1964 . GENETICS SOC JAPAN NATL INST GENETICS, YATA, MISHIMA, SHIZUOKA 411, JAPAN: 340-&. Bhattacharya B: Cytological studies on some Indian members of Commelinaceae. Cytologia 1975, 40(2):285-299. Alam N: Trends of Chromosome evolution in family Commelinaceae. The Nucleus 1984, 27:231-241. Eksomtramage L, Sirirugsa P, Sawangchote P, Jornead S, Saknimit T, Leeratiwong C: Chromosome number of some monocot species from Ton-Nga-Chang Wildlife Sanctuary, southern Thailand. Thai Forest Bulletin (Botany) 2001(29):63-71. Fujishima H: Karyotypic diversity of Commelina benghalensis L.(Commelinaceae). Chromosome science 2007, 10(2):43-53. Fujishima H: Karyotypic diversity of Commelina diffusa Burm.(Commelinaceae). Chromosome science 2007, 10(1):21-28. Morton JK: The Commelinaceae of West Africa: a biosystematic survey. Bot J Linn Soc 1967, 60(382):167-221. Kammathy R, Rolla SR: Notes on Indian Commelinaceae-II: Cytological Observation. Nelumbo 1961, 3(2):167-169. RAGHAVAN RS: Cytological observation on the Indian species of Commelinaceae. Current Sci 1961, 30:310-311. Morton J: Cytotaxonomic studies on the Gold Coast species of the genus Commelina Linn. Bot J Linn Soc 1956, 55(361):507-531. Shigenobu Y, Kobori M: Karyotype variation of Commelina benghalensis correlated with habitats in Kochi Prefecture, Japan. Chromosome science 1997, 1(2):83-87. Oziegbe M, Eludini PO: Karyotypic Studies of Commelina benghalensis variety benghalensis and C. forskalaei (Commelinaceae) from Nigeria. Cytologia 2013, 78(2):151-156. Panigraphi G: Cytotaxonomic studies in certain species of Commelina Linn. in eastern India. J Indian Bot Soc 1964, 43:294-310. Gowda SR, Gawde HM, Hyderi A, Savitha MR, Krishnamurthy B, Karat SC, Doddaiah N, Patel ZM, Ramachandra NB: Chromosomal anomalies and congenital heart disease in Mysore, South India. Int J Hum Genet 2010, 10(1-3):131-139. Carter S, Cutler D, Reynold T, Brandham P: A multidisciplinary approach to a revision of the Aloe somaliensis complex (Liliaceae). Kew bulletin 1984:611-633. Dagne K: Karyotypes, C-banding and nucleolar numbers inGuizotia (Compositae). Plant Syst Evol 1995, 195(1-2):121-135. Dagne K, HENEEN WK: The karyotype and nucleoli of Guizotia abyssinica (Compositae). Hereditas 1992, 117(1):73-83. Levan A: Nomenclature for centromeric position on chromosomes. Hereditas 1964, 52:201-220. Zarco CR: A new method for estimating karyotype asymmetry. Taxon 1986, 35(3):526-530. Lavania U, Srivastava S: Quantitative delineation of karyotype variation in Papaver as a measure of phylogenetic differentiation and origin. Curr Sci 1999:429-435. Gunjan K, Roy BK: Karyotype studies in dominant species of Aloe from eastern India. Caryologia 2010, 63(1):41-49. 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Also discoverable on Platform About Our Team In Review Editorial Policies Advisory Board Help Center Resources Author Services Accessibility API Access RSS feed Manage Cookie Preferences © Research Square 2026 | ISSN 2693-5015 (online) Privacy Policy Terms of Service Do Not Sell My Personal Information {"props":{"pageProps":{"initialData":{"identity":"rs-100645","acceptedTermsAndConditions":true,"allowDirectSubmit":true,"archivedVersions":[],"articleType":"Research","associatedPublications":[],"authors":[{"id":4194241,"identity":"cae2df35-2c7a-47fc-a844-877da29d51b4","order_by":0,"name":"Samuel Gebrechristos Gebretsadkan","email":"data:image/png;base64,iVBORw0KGgoAAAANSUhEUgAAAZAAAAAyAQMAAABI0h/eAAAABlBMVEX///8AAABVwtN+AAAACXBIWXMAAA7EAAAOxAGVKw4bAAAAy0lEQVRIiWNgGAWjYLACxgYGxn4QI6GAFC0zG0BaDEjRsuEAiEWMFoPj7Q8/3dxhI7v5/OrEDw8MGOT5xQ4Q0HLmjLF07pk042033m6WADrMcObsBAJabuQwSOe2HU7cduPsBpCWBIPbhLTcf/74N0jL5hlnN/8gTssNBjOwLRv4e7cRZ4vkmRwza5BfZtzg3WaRYCBB2C98x48/vp0LDLH+/rObb/6osJHnlyagReEAjCUBVimBXzkIyDfAWPwHcKsaBaNgFIyCkQ0ApUtNf2MN6ZAAAAAASUVORK5CYII=","orcid":"https://orcid.org/0000-0002-6646-7303","institution":"Aksum University","correspondingAuthor":true,"submittingAuthor":false,"prefix":"","firstName":"Samuel","middleName":"Gebrechristos","lastName":"Gebretsadkan","suffix":""},{"id":4194242,"identity":"1fe43026-4c72-43aa-ade9-0352a6fd25e8","order_by":1,"name":"Gebrehiwet Kiros","email":"","orcid":"","institution":"Aksum University","correspondingAuthor":false,"submittingAuthor":false,"prefix":"","firstName":"Gebrehiwet","middleName":"","lastName":"Kiros","suffix":""},{"id":4194243,"identity":"daec26ab-b9df-4352-b03c-87fa4f9f49f9","order_by":2,"name":"Bahlibi Gebreabzgi","email":"","orcid":"","institution":"Aksum University","correspondingAuthor":false,"submittingAuthor":false,"prefix":"","firstName":"Bahlibi","middleName":"","lastName":"Gebreabzgi","suffix":""},{"id":4194244,"identity":"8de07975-f6c0-4c47-b0a5-7351cd8aaafc","order_by":3,"name":"Destalem Gebremichael","email":"","orcid":"","institution":"Tigray Agricultural Research Institute","correspondingAuthor":false,"submittingAuthor":false,"prefix":"","firstName":"Destalem","middleName":"","lastName":"Gebremichael","suffix":""},{"id":4194245,"identity":"b104395c-3d04-4cf8-a83d-6daf60becea3","order_by":4,"name":"Guesh Mulaw","email":"","orcid":"","institution":"Aksum University","correspondingAuthor":false,"submittingAuthor":false,"prefix":"","firstName":"Guesh","middleName":"","lastName":"Mulaw","suffix":""}],"badges":[],"createdAt":"2020-10-30 13:01:41","currentVersionCode":1,"declarations":"","doi":"10.21203/rs.3.rs-100645/v1","doiUrl":"https://doi.org/10.21203/rs.3.rs-100645/v1","draftVersion":[],"editorialEvents":[],"editorialNote":"","failedWorkflow":false,"files":[{"id":3376445,"identity":"8d8e2018-ff98-4e34-a841-d949d752a915","added_by":"auto","created_at":"2020-11-04 15:18:12","extension":"jpg","order_by":1,"title":"Figure 1","display":"","copyAsset":false,"role":"figure","size":106752,"visible":true,"origin":"","legend":"Somatic metaphase spread and karyotype of the genus Commelina L.; C. africana (A \u0026 B), C. benghalensis (C \u0026D), C. diffusa (Ginchi) (E \u0026 F), C. diffusa (Jimma) (G \u0026H), C. subulata (I\u0026J); A, C, E, G \u0026 I reveal the metaphase spread where as the remaining images are karyotypes; horizontal bar indicates the scale at 5 µm for C. benghalensis \u0026 C. subulata where as the scale bar for C. africana, C. diffusa (Ginchi) \u0026 C. diffusa (Jimma) is 10 µm","description":"","filename":"Figure1.JPG","url":"https://assets-eu.researchsquare.com/files/rs-100645/v1/b4018ed1fb8845471409e5ed.JPG"},{"id":3376441,"identity":"8f6b1cbb-e3b1-4a47-9a76-72d4251fdd06","added_by":"auto","created_at":"2020-11-04 15:18:05","extension":"jpg","order_by":1,"title":"Figure 1","display":"","copyAsset":false,"role":"figure","size":106752,"visible":true,"origin":"","legend":"Somatic metaphase spread and karyotype of the genus Commelina L.; C. africana (A \u0026 B), C. benghalensis (C \u0026D), C. diffusa (Ginchi) (E \u0026 F), C. diffusa (Jimma) (G \u0026H), C. subulata (I\u0026J); A, C, E, G \u0026 I reveal the metaphase spread where as the remaining images are karyotypes; horizontal bar indicates the scale at 5 µm for C. benghalensis \u0026 C. subulata where as the scale bar for C. africana, C. diffusa (Ginchi) \u0026 C. diffusa (Jimma) is 10 µm","description":"","filename":"Figure1.JPG","url":"https://assets-eu.researchsquare.com/files/rs-100645/v1/25ac13e08cd49bdda65719bc.JPG"},{"id":3376446,"identity":"88c29111-2401-435c-9a05-4fdffec829d0","added_by":"auto","created_at":"2020-11-04 15:18:12","extension":"jpg","order_by":2,"title":"Figure 2","display":"","copyAsset":false,"role":"figure","size":120523,"visible":true,"origin":"","legend":"NORs and interphase nucleoli of Commelina L. A. two satellites of C. africana B. Three nucleoli of C. africana C. five NORs of C. diffusa from Ginchi; D, E, F. \u0026 G indicates nucleoli of C. diffusa (Ginchi), C. benghalensis, C. diffusa (jimma) and C. subulata respectively. Arraws indicate the specific position of satellites","description":"","filename":"Figure2.JPG","url":"https://assets-eu.researchsquare.com/files/rs-100645/v1/36e6a4a7a848410ef3aa283a.JPG"},{"id":3376442,"identity":"9af82986-ad3b-4629-9ead-f2f71436c356","added_by":"auto","created_at":"2020-11-04 15:18:06","extension":"jpg","order_by":2,"title":"Figure 2","display":"","copyAsset":false,"role":"figure","size":120523,"visible":true,"origin":"","legend":"NORs and interphase nucleoli of Commelina L. A. two satellites of C. africana B. Three nucleoli of C. africana C. five NORs of C. diffusa from Ginchi; D, E, F. \u0026 G indicates nucleoli of C. diffusa (Ginchi), C. benghalensis, C. diffusa (jimma) and C. subulata respectively. Arraws indicate the specific position of satellites","description":"","filename":"Figure2.JPG","url":"https://assets-eu.researchsquare.com/files/rs-100645/v1/fb6671f2cccc2f9b9599dfe1.JPG"},{"id":13609127,"identity":"f28501ae-0513-4ccf-b4dd-7634e16dec6f","added_by":"auto","created_at":"2021-09-17 06:18:57","extension":"pdf","order_by":0,"title":"","display":"","copyAsset":false,"role":"manuscript-pdf","size":632045,"visible":true,"origin":"","legend":"","description":"","filename":"manuscript.pdf","url":"https://assets-eu.researchsquare.com/files/rs-100645/v1/5e80f5b3-5ed0-47cb-994e-f7ea6d73c702.pdf"}],"financialInterests":"","formattedTitle":"\u003cp\u003eKaryo-Morphology and Nucleoli Analysis of Commelina L. (Commelinaceae) from Ethiopia\u003c/p\u003e","fulltext":[{"header":"Background","content":"\u003cp\u003eThe family Commelinaceae, commonly known as the spiderwort, is known to possess 41 genera and 650 species throughout the world [\u003ca href=\"#_ENREF_1\"\u003e1\u003c/a\u003e], with main distribution in tropics and subtropics that extends into northern temperate regions. The family is represented by 9 genera and 56 species in Ethiopia [\u003ca href=\"#_ENREF_2\"\u003e2\u003c/a\u003e].With about 100 species, \u003cem\u003eCommelina \u003c/em\u003eis the largest genus of Commelinaceae in Africa. At least 65 species occur in the combined areas of the Flora of Tropical East Africa (Kenya, Uganda, and Tanzania) and Flora Zambesiaca (Malawi, Mozambique, Zambia, Zimbabwe, and Botswana) [\u003ca href=\"#_ENREF_3\"\u003e3\u003c/a\u003e, \u003ca href=\"#_ENREF_4\"\u003e4\u003c/a\u003e]. About 19 species have been identified in Ethiopia and Eritrea including 2 un-described species that are endemic to Ethiopia [\u003ca href=\"#_ENREF_5\"\u003e5\u003c/a\u003e].\u003c/p\u003e\n\u003cp\u003eThe genus \u003cem\u003eCommelina \u003c/em\u003ehas several medicinal significances. \u003cem\u003eC. benghalensis \u003c/em\u003eis used to treat bed sores, breast sores and pimples in Pakistan [\u003ca href=\"#_ENREF_6\"\u003e6\u003c/a\u003e]. In East Africa, the sap of \u003cem\u003eC. benghalensis \u003c/em\u003eleaves and stems is used to treat ophthalmia, infertility in women, leprosy, sore throat and burns, and the liquid contained in the flowering spathe is used to treat eye complaints in Zanzibar [\u003ca href=\"#_ENREF_7\"\u003e7\u003c/a\u003e]. People from Nepal use a paste derived from the plant to treat burns, and indigestion with a juice produced from the roots [\u003ca href=\"#_ENREF_8\"\u003e8\u003c/a\u003e]. In China, \u003cem\u003eC. diffusa \u003c/em\u003eis used as a medicinal herb with febrifugal and diuretic effects [\u003ca href=\"#_ENREF_9\"\u003e9\u003c/a\u003e].\u003c/p\u003e\n\u003cp\u003e\u003cem\u003eC. benghalensis\u003c/em\u003e is known for its medicinal purposes as they have already been reported in treating hypertension, burns, leprosy, sore throats, cataplasm, and wound healing [\u003ca href=\"#_ENREF_10\"\u003e10\u003c/a\u003e]. Numerous compounds have been identified from the vegetative and flower parts of \u003cem\u003eCommelina benghalensis \u003c/em\u003eincluding noctacosanol, n-triocotanol, stigma-sterol, compesterol and hydrocyanic acid [\u003ca href=\"#_ENREF_11\"\u003e11\u003c/a\u003e]. Phyto-chemical screening also revealed the presence of many secondary metabolites like phlobatannins, carbohydrates, tannins, glycosides, volatile oils, resins, balsams, flavonoids and saponins [\u003ca href=\"#_ENREF_12\"\u003e12\u003c/a\u003e]. Presence of flavonoids, for example, indicates the plant might have an antioxidant, anti-allergic, anti-inflammatory, anti- microbial or anti- cancer activity [\u003ca href=\"#_ENREF_13\"\u003e13\u003c/a\u003e]. In addition, \u003cem\u003eC. benghalensis \u003c/em\u003ehas analgestic action that proves the folkloric use in pain management [\u003ca href=\"#_ENREF_14\"\u003e14\u003c/a\u003e].\u003c/p\u003e\n\u003cp\u003eAlthough several medicinal and economic uses of the family Commelinaceae are known, little has been done in cytological analysis in Ethiopia. Previous research done\u0026nbsp;only focused on chromosome number of four species included in this study based on meiosis and voucher specimen were taken only from the Harar Province except for \u003cem\u003eCyanotis barbata \u003c/em\u003eD. Don which was from Debrezeit [\u003ca href=\"#_ENREF_15\"\u003e15\u003c/a\u003e]. The present study was focused on chromosome number, karyotype analysis and silver staining. Here the chromosome number, ploidy level, karyotype and number of nucleoli organizing region (NORs) of these species of Commelinaceae are described. Such study can help to expand the current cytological knowledge of the Ethiopian Commelinaceae with further contribution for phylogenetic and biosystematics research.\u003c/p\u003e"},{"header":"Results","content":"\u003cp\u003eThe \u003cem\u003eCommelina \u003c/em\u003especies contain metacentric, sub-metacentric and sub-telocentric chromosomes with variation in number and morphology (Table 1).\u003c/p\u003e\n\u003ch3\u003eC. africana\u003c/h3\u003e\n\u003cp\u003eSomatic chromosomes observed from more than five intact cells showed that \u003cem\u003eC. africana \u003c/em\u003ecollected from Addis Ababa contained a diploid number of 2\u003cem\u003en\u003c/em\u003e=2\u003cem\u003ex\u003c/em\u003e= 30. Because of its better chromosome spread and clear chromosome morphology a cell with 29 chromosome number is shown in Figure 1A and a karyotype constructed from same cell in Figure 1B. This variant of the species consisted of six pairs of metacentric chromosomes (pairs 1 to 6), five pairs of sub- metacentic chromosomes (pair numbers 7 to 11) and four sub-telocentrics (pairs 12 to 15) (Figure 1B). The karyotype formula is, therefore, 12m + 10sm + 8 st (Table 1). According to Stebbins karyotype classification, this species belongs to 2A type with asymmetry index of 65.19 (Table 1). Small satellites were also observed at the tip of the short arm of a pair of chromosomes (Figure 2A) and these correspond to the 8\u003csup\u003eth\u003c/sup\u003e pair on the karyotype (Figure 1B). Figure 2B shows three interphase nucleoli, from which it can be assumed that the maximum number of nucleoli and so chromosome satellites for this species is four.\u003c/p\u003e\n\u003cp\u003eLength wise measurement of metaphase chromosomes revealed a value range from the smallest 4.53\u0026micro;m to largest 7.5\u0026micro;m (Table 1). Total length of whole chromosome complement (2\u003cem\u003en\u003c/em\u003e) was 176.72\u0026micro;m. The chromosomes were medium in size with an average length and asymmetry of 5.89\u0026micro;m and 1.87, respectively.\u003c/p\u003e\n\u003ch3\u003eC. benghalensis\u003c/h3\u003e\n\u003cp\u003eAccording to somatic chromosome analysis of the root tips, this species possesses 2\u003cem\u003en\u003c/em\u003e=6\u003cem\u003ex\u003c/em\u003e=66 (Figure 1C and Table 1). The karyotpe constructed based on calculated arm ratio and chromosome size revealed the presence of eighteen metacentric chromosome pairs (pair numbers 1, 3, 4, 5, 8, 11, 14, 15, 16, 18, 22, 24, 25, 29, 30, 31, 32, 33), twelve sub-metacentrics (pair numbers 6, 7, 9, 10, 12, 17, 19, 20, 21, 23, 27, 28) and 3 sub-telocentric pairs (pairs 2, 13, 26 ) (Figure 1D). The karyotype formula was, therefore, 36m + 24sm + 6st. As to Stebbins method of classification, \u003cem\u003eC. benghalensis \u003c/em\u003epossesses asymmetry index of 62.58 with 2B in karyotype asymmetry (Table 1). A maximum of five nucleoli were observed through silver staining but no satellites were observed (Figure 1E). Two of the nucleoli are much larger than the other three nucleoli. Each of the larger nucleoli might have resulted from fusion of two or more nucleoli. It can be inferred that six or more chromosome satellites are present in this species.\u003c/p\u003e\n\u003cp\u003eThe chromosome lengths differ from the smallest 2.5\u0026micro;m and gradually increased up to the largest 5.625\u0026micro;m (Table 1). The total length of whole chromosome complement was about 267.935\u0026micro;m (Table 1). The total length of the long arms of the diploid set was 167.675\u0026micro;m and that of short arms for diploid set was 100.26\u0026micro;m with their ratio generating an average karyotype asymmetry of 1.672. Generally, \u003cem\u003eC. benghalensis \u003c/em\u003e(Addis Ababa) has predominantly metacentric and sub-metacentric type of chromosomes. The chromosomes were short to medium in size with an average length of 4.06\u0026micro;m.\u003c/p\u003e\n\u003cp\u003e\u003cstrong\u003e\u003cem\u003eC. diffusa \u003c/em\u003e\u003c/strong\u003e\u003cstrong\u003efrom Ginchi\u003c/strong\u003e\u003c/p\u003e\n\u003cp\u003eThe variant of \u003cem\u003eC. diffusa \u003c/em\u003ecollected from Ginchi possessed 2\u003cem\u003en\u003c/em\u003e= 66 (Table 1 and Figure 1E). Based on centromeric ratio and chromosome length measurements , the karyotype (Figure 1F) consisted of fourteen metacentric pairs (pair numbers 1, 4, 6, 15, 16, 20, 21, 23, 25, 26, 30, 31, 32, 33), thirteen sub-metacentrics (pairs 2, 3, 5, 7, 8, 9, 10, 13, 17, 19, 24, 27, 28) and six sub-telocentrics (pairs 11, 12, 14, 18, 22, 29) with a formula of 28m + 26sm + 12st. The asymmetry index is 65.56 and belongs to 2B type of Stebbins karyotype classification (Table 1).\u003c/p\u003e\n\u003cp\u003eAs shown in Figure 2(C and D), a total of six satellites and four nucleoli were observed from chromosome preparation and silver staining, respectively. Size of satellites varies with one pair being larger than the other two.\u003c/p\u003e\n\u003cp\u003e\u003cem\u003eC. diffusa \u003c/em\u003e(Ginchi) has comparatively larger chromosomes than the other \u003cem\u003eCommelina \u003c/em\u003especies included in this study with a length range from smallest 4.375\u0026micro;m to largest 11.25\u0026micro;m and total length of whole chromosome complement 507.53\u0026micro;m (Table 1). The total length of long arms and short arms for the diploid set was 332.745\u0026micro;m and 174.785\u0026micro;m respectively (Table 1). The average karyotype asymmetry is 1.904 and on the average the chromosomes can be classified under sub-metacentric. The chromosomes were medium to large in size with an average length of 7.69\u0026micro;m.\u003c/p\u003e\n\u003cp\u003e\u003cem\u003eC. diffusa \u003c/em\u003e(Ginchi) has intra-chromosomal and inter-chromosomal asymmetry indices of 0.435 and 0.17035 respectively (Table 1). This variant also has a coefficient of variation in chromosome length of (17.035), ratio of centromeric gradient (53.0163) and dispersion index (9.031) (Table 1).\u003c/p\u003e\n\u003cp\u003e\u003cstrong\u003eTable 1 Cytological analysis of Commelina L.\u003c/strong\u003e\u003c/p\u003e\n\u003ctable border=\"1\" width=\"0\"\u003e\n\u003ctbody\u003e\n\u003ctr\u003e\n\u003ctd rowspan=\"2\" width=\"98\"\u003e\n\u003cp\u003e\u003cstrong\u003eSpecies\u003c/strong\u003e\u003c/p\u003e\n\u003c/td\u003e\n\u003ctd rowspan=\"2\" width=\"69\"\u003e\n\u003cp\u003e\u003cstrong\u003eChromosome \u003c/strong\u003e\u003cstrong\u003elength (\u0026micro;m)\u003c/strong\u003e\u003c/p\u003e\n\u003cp\u003e\u003cstrong\u003eShortest: longest\u003c/strong\u003e\u003c/p\u003e\n\u003c/td\u003e\n\u003ctd rowspan=\"2\" width=\"50\"\u003e\n\u003cp\u003e\u003cstrong\u003eACL (\u0026micro;m)\u003c/strong\u003e\u003c/p\u003e\n\u003c/td\u003e\n\u003ctd rowspan=\"2\" width=\"42\"\u003e\n\u003cp\u003e\u003cstrong\u003e2\u003cem\u003en\u003c/em\u003e\u003c/strong\u003e\u003c/p\u003e\n\u003c/td\u003e\n\u003ctd rowspan=\"2\" width=\"50\"\u003e\n\u003cp\u003e\u003cstrong\u003eBasic number(\u003cem\u003ex\u003c/em\u003e)\u003c/strong\u003e\u003c/p\u003e\n\u003c/td\u003e\n\u003ctd rowspan=\"2\" width=\"42\"\u003e\n\u003cp\u003e\u003cstrong\u003ePloidy level\u003c/strong\u003e\u003c/p\u003e\n\u003c/td\u003e\n\u003ctd rowspan=\"2\" width=\"50\"\u003e\n\u003cp\u003e\u003cstrong\u003eA\u003csub\u003e1\u003c/sub\u003e\u003c/strong\u003e\u003c/p\u003e\n\u003c/td\u003e\n\u003ctd rowspan=\"2\" width=\"50\"\u003e\n\u003cp\u003e\u003cstrong\u003eA\u003csub\u003e2\u003c/sub\u003e\u003c/strong\u003e\u003c/p\u003e\n\u003c/td\u003e\n\u003ctd rowspan=\"2\" width=\"50\"\u003e\n\u003cp\u003e\u003cstrong\u003eC\u003csub\u003ev\u003c/sub\u003e\u003c/strong\u003e\u003c/p\u003e\n\u003c/td\u003e\n\u003ctd rowspan=\"2\" width=\"50\"\u003e\n\u003cp\u003e\u003cstrong\u003eC\u003csub\u003eG\u003c/sub\u003e\u003c/strong\u003e\u003c/p\u003e\n\u003c/td\u003e\n\u003ctd rowspan=\"2\" width=\"50\"\u003e\n\u003cp\u003e\u003cstrong\u003eDI\u003c/strong\u003e\u003c/p\u003e\n\u003c/td\u003e\n\u003ctd rowspan=\"2\" width=\"69\"\u003e\n\u003cp\u003e\u003cstrong\u003eKaryotype Formula\u003c/strong\u003e\u003c/p\u003e\n\u003c/td\u003e\n\u003ctd rowspan=\"2\" width=\"42\"\u003e\n\u003cp\u003e\u003cstrong\u003eNumber of metaphase plates\u003c/strong\u003e\u003c/p\u003e\n\u003c/td\u003e\n\u003ctd rowspan=\"2\" width=\"55\"\u003e\n\u003cp\u003e\u003cstrong\u003eAsymmetry index\u003c/strong\u003e\u003c/p\u003e\n\u003c/td\u003e\n\u003ctd colspan=\"3\" width=\"133\"\u003e\n\u003cp\u003e\u003cstrong\u003eStebbins asymmetry\u003c/strong\u003e\u003c/p\u003e\n\u003c/td\u003e\n\u003c/tr\u003e\n\u003ctr\u003e\n\u003ctd width=\"48\"\u003e\n\u003cp\u003e\u003cstrong\u003eK\u003c/strong\u003e\u003c/p\u003e\n\u003c/td\u003e\n\u003ctd width=\"48\"\u003e\n\u003cp\u003e\u003cstrong\u003eL\u003c/strong\u003e\u003c/p\u003e\n\u003c/td\u003e\n\u003ctd width=\"36\"\u003e\n\u003cp\u003e\u003cstrong\u003eM\u003c/strong\u003e\u003c/p\u003e\n\u003c/td\u003e\n\u003c/tr\u003e\n\u003ctr\u003e\n\u003ctd width=\"98\"\u003e\n\u003cp\u003e\u003cem\u003eC. africana \u003c/em\u003e\u003c/p\u003e\n\u003c/td\u003e\n\u003ctd width=\"69\"\u003e\n\u003cp\u003e4.53 \u0026ndash; 7.5\u003c/p\u003e\n\u003c/td\u003e\n\u003ctd width=\"50\"\u003e\n\u003cp\u003e5.89\u003c/p\u003e\n\u003c/td\u003e\n\u003ctd width=\"42\"\u003e\n\u003cp\u003e30\u003c/p\u003e\n\u003c/td\u003e\n\u003ctd width=\"50\"\u003e\n\u003cp\u003e15\u003c/p\u003e\n\u003c/td\u003e\n\u003ctd width=\"42\"\u003e\n\u003cp\u003e2x\u003c/p\u003e\n\u003c/td\u003e\n\u003ctd width=\"50\"\u003e\n\u003cp\u003e0.4345\u003c/p\u003e\n\u003c/td\u003e\n\u003ctd width=\"50\"\u003e\n\u003cp\u003e0.1498\u003c/p\u003e\n\u003c/td\u003e\n\u003ctd width=\"50\"\u003e\n\u003cp\u003e14.98\u003c/p\u003e\n\u003c/td\u003e\n\u003ctd width=\"50\"\u003e\n\u003cp\u003e55.32\u003c/p\u003e\n\u003c/td\u003e\n\u003ctd width=\"50\"\u003e\n\u003cp\u003e8.29\u003c/p\u003e\n\u003c/td\u003e\n\u003ctd width=\"69\"\u003e\n\u003cp\u003e12m + 10sm + 8st\u003c/p\u003e\n\u003c/td\u003e\n\u003ctd width=\"42\"\u003e\n\u003cp\u003e10\u003c/p\u003e\n\u003c/td\u003e\n\u003ctd width=\"55\"\u003e\n\u003cp\u003e65.19\u003c/p\u003e\n\u003c/td\u003e\n\u003ctd width=\"48\"\u003e\n\u003cp\u003e0.43\u003c/p\u003e\n\u003c/td\u003e\n\u003ctd width=\"48\"\u003e\n\u003cp\u003e1.67\u003c/p\u003e\n\u003c/td\u003e\n\u003ctd width=\"36\"\u003e\n\u003cp\u003e2A\u003c/p\u003e\n\u003c/td\u003e\n\u003c/tr\u003e\n\u003ctr\u003e\n\u003ctd width=\"98\"\u003e\n\u003cp\u003e\u003cem\u003eC. benghalensis\u003c/em\u003e\u003c/p\u003e\n\u003c/td\u003e\n\u003ctd width=\"69\"\u003e\n\u003cp\u003e2.5 \u0026ndash; 5.625\u003c/p\u003e\n\u003c/td\u003e\n\u003ctd width=\"50\"\u003e\n\u003cp\u003e4.06\u003c/p\u003e\n\u003c/td\u003e\n\u003ctd width=\"42\"\u003e\n\u003cp\u003e66\u003c/p\u003e\n\u003c/td\u003e\n\u003ctd width=\"50\"\u003e\n\u003cp\u003e11\u003c/p\u003e\n\u003c/td\u003e\n\u003ctd width=\"42\"\u003e\n\u003cp\u003e6\u003cem\u003ex\u003c/em\u003e\u003c/p\u003e\n\u003c/td\u003e\n\u003ctd width=\"50\"\u003e\n\u003cp\u003e0.3715\u003c/p\u003e\n\u003c/td\u003e\n\u003ctd width=\"50\"\u003e\n\u003cp\u003e0.1582\u003c/p\u003e\n\u003c/td\u003e\n\u003ctd width=\"50\"\u003e\n\u003cp\u003e15.82\u003c/p\u003e\n\u003c/td\u003e\n\u003ctd width=\"50\"\u003e\n\u003cp\u003e57.831\u003c/p\u003e\n\u003c/td\u003e\n\u003ctd width=\"50\"\u003e\n\u003cp\u003e0.3715\u003c/p\u003e\n\u003c/td\u003e\n\u003ctd width=\"69\"\u003e\n\u003cp\u003e36m + 24sm + 6st\u003c/p\u003e\n\u003c/td\u003e\n\u003ctd width=\"42\"\u003e\n\u003cp\u003e4\u003c/p\u003e\n\u003c/td\u003e\n\u003ctd width=\"55\"\u003e\n\u003cp\u003e62.58\u003c/p\u003e\n\u003c/td\u003e\n\u003ctd width=\"48\"\u003e\n\u003cp\u003e0.35\u003c/p\u003e\n\u003c/td\u003e\n\u003ctd width=\"48\"\u003e\n\u003cp\u003e2.25\u003c/p\u003e\n\u003c/td\u003e\n\u003ctd width=\"36\"\u003e\n\u003cp\u003e2B\u003c/p\u003e\n\u003c/td\u003e\n\u003c/tr\u003e\n\u003ctr\u003e\n\u003ctd width=\"98\"\u003e\n\u003cp\u003e\u003cem\u003eC.diffusa \u003c/em\u003e(Ginchi)\u003c/p\u003e\n\u003c/td\u003e\n\u003ctd width=\"69\"\u003e\n\u003cp\u003e4.375 \u0026ndash; 11.25\u003c/p\u003e\n\u003c/td\u003e\n\u003ctd width=\"50\"\u003e\n\u003cp\u003e7.69\u003c/p\u003e\n\u003c/td\u003e\n\u003ctd width=\"42\"\u003e\n\u003cp\u003e66\u003c/p\u003e\n\u003c/td\u003e\n\u003ctd width=\"50\"\u003e\n\u003cp\u003e-\u003c/p\u003e\n\u003c/td\u003e\n\u003ctd width=\"42\"\u003e\n\u003cp\u003e-\u003c/p\u003e\n\u003c/td\u003e\n\u003ctd width=\"50\"\u003e\n\u003cp\u003e0.435\u003c/p\u003e\n\u003c/td\u003e\n\u003ctd width=\"50\"\u003e\n\u003cp\u003e0.17035\u003c/p\u003e\n\u003c/td\u003e\n\u003ctd width=\"50\"\u003e\n\u003cp\u003e17.035\u003c/p\u003e\n\u003c/td\u003e\n\u003ctd width=\"50\"\u003e\n\u003cp\u003e53.0163\u003c/p\u003e\n\u003c/td\u003e\n\u003ctd width=\"50\"\u003e\n\u003cp\u003e9.031\u003c/p\u003e\n\u003c/td\u003e\n\u003ctd width=\"69\"\u003e\n\u003cp\u003e28m + 26sm + 12st\u003c/p\u003e\n\u003c/td\u003e\n\u003ctd width=\"42\"\u003e\n\u003cp\u003e5\u003c/p\u003e\n\u003c/td\u003e\n\u003ctd width=\"55\"\u003e\n\u003cp\u003e65.56\u003c/p\u003e\n\u003c/td\u003e\n\u003ctd width=\"48\"\u003e\n\u003cp\u003e0.5\u003c/p\u003e\n\u003c/td\u003e\n\u003ctd width=\"48\"\u003e\n\u003cp\u003e2.57\u003c/p\u003e\n\u003c/td\u003e\n\u003ctd width=\"36\"\u003e\n\u003cp\u003e2B\u003c/p\u003e\n\u003c/td\u003e\n\u003c/tr\u003e\n\u003ctr\u003e\n\u003ctd width=\"98\"\u003e\n\u003cp\u003e\u003cem\u003eC.diffusa \u003c/em\u003e(Jimma)\u003c/p\u003e\n\u003c/td\u003e\n\u003ctd width=\"69\"\u003e\n\u003cp\u003e2.5 \u0026ndash; 4.58\u003c/p\u003e\n\u003c/td\u003e\n\u003ctd width=\"50\"\u003e\n\u003cp\u003e3.47\u003c/p\u003e\n\u003c/td\u003e\n\u003ctd width=\"42\"\u003e\n\u003cp\u003e30\u003c/p\u003e\n\u003c/td\u003e\n\u003ctd width=\"50\"\u003e\n\u003cp\u003e15\u003c/p\u003e\n\u003c/td\u003e\n\u003ctd width=\"42\"\u003e\n\u003cp\u003e2\u003cem\u003ex\u003c/em\u003e\u003c/p\u003e\n\u003c/td\u003e\n\u003ctd width=\"50\"\u003e\n\u003cp\u003e0.76717\u003c/p\u003e\n\u003c/td\u003e\n\u003ctd width=\"50\"\u003e\n\u003cp\u003e0.13771\u003c/p\u003e\n\u003c/td\u003e\n\u003ctd width=\"50\"\u003e\n\u003cp\u003e13.771\u003c/p\u003e\n\u003c/td\u003e\n\u003ctd width=\"50\"\u003e\n\u003cp\u003e39.32\u003c/p\u003e\n\u003c/td\u003e\n\u003ctd width=\"50\"\u003e\n\u003cp\u003e5.415\u003c/p\u003e\n\u003c/td\u003e\n\u003ctd width=\"69\"\u003e\n\u003cp\u003e10m + 8sm + 12st\u003c/p\u003e\n\u003c/td\u003e\n\u003ctd width=\"42\"\u003e\n\u003cp\u003e10\u003c/p\u003e\n\u003c/td\u003e\n\u003ctd width=\"55\"\u003e\n\u003cp\u003e69.497\u003c/p\u003e\n\u003c/td\u003e\n\u003ctd width=\"48\"\u003e\n\u003cp\u003e0.67\u003c/p\u003e\n\u003c/td\u003e\n\u003ctd width=\"48\"\u003e\n\u003cp\u003e1.832\u003c/p\u003e\n\u003c/td\u003e\n\u003ctd width=\"36\"\u003e\n\u003cp\u003e3A\u003c/p\u003e\n\u003c/td\u003e\n\u003c/tr\u003e\n\u003ctr\u003e\n\u003ctd width=\"98\"\u003e\n\u003cp\u003e\u003cem\u003eC. subulata\u003c/em\u003e\u003c/p\u003e\n\u003c/td\u003e\n\u003ctd width=\"69\"\u003e\n\u003cp\u003e1.923 \u0026ndash; 3.654\u003c/p\u003e\n\u003c/td\u003e\n\u003ctd width=\"50\"\u003e\n\u003cp\u003e2.77\u003c/p\u003e\n\u003c/td\u003e\n\u003ctd width=\"42\"\u003e\n\u003cp\u003e30\u003c/p\u003e\n\u003c/td\u003e\n\u003ctd width=\"50\"\u003e\n\u003cp\u003e15\u003c/p\u003e\n\u003c/td\u003e\n\u003ctd width=\"42\"\u003e\n\u003cp\u003e2\u003cem\u003ex\u003c/em\u003e\u003c/p\u003e\n\u003c/td\u003e\n\u003ctd width=\"50\"\u003e\n\u003cp\u003e0.3523\u003c/p\u003e\n\u003c/td\u003e\n\u003ctd width=\"50\"\u003e\n\u003cp\u003e0.1891\u003c/p\u003e\n\u003c/td\u003e\n\u003ctd width=\"50\"\u003e\n\u003cp\u003e18.91\u003c/p\u003e\n\u003c/td\u003e\n\u003ctd width=\"50\"\u003e\n\u003cp\u003e67.45\u003c/p\u003e\n\u003c/td\u003e\n\u003ctd width=\"50\"\u003e\n\u003cp\u003e0.3523\u003c/p\u003e\n\u003c/td\u003e\n\u003ctd width=\"69\"\u003e\n\u003cp\u003e18m + 10sm + 2st\u003c/p\u003e\n\u003c/td\u003e\n\u003ctd width=\"42\"\u003e\n\u003cp\u003e4\u003c/p\u003e\n\u003c/td\u003e\n\u003ctd width=\"55\"\u003e\n\u003cp\u003e60.232\u003c/p\u003e\n\u003c/td\u003e\n\u003ctd width=\"48\"\u003e\n\u003cp\u003e0.267\u003c/p\u003e\n\u003c/td\u003e\n\u003ctd width=\"48\"\u003e\n\u003cp\u003e1.9\u003c/p\u003e\n\u003c/td\u003e\n\u003ctd width=\"36\"\u003e\n\u003cp\u003e2A\u003c/p\u003e\n\u003c/td\u003e\n\u003c/tr\u003e\n\u003c/tbody\u003e\n\u003c/table\u003e\n\u003cp\u003e\u003cstrong\u003eACL:\u003c/strong\u003e average chromosome length; 2\u003cem\u003en\u003c/em\u003e: somatic chromosome number; \u003cstrong\u003eA1: \u003c/strong\u003eintra-chromosomal asymmetry index; \u003cstrong\u003eA2: \u003c/strong\u003einter-chromosomal asymmetry index; \u003cstrong\u003eC\u003csub\u003ev\u003c/sub\u003e: \u003c/strong\u003eCoefficient of variation of chromosome length; \u003cstrong\u003eC\u003csub\u003eG\u003c/sub\u003e: \u003c/strong\u003eratio of centromeric gradient; \u003cstrong\u003eDI: \u003c/strong\u003edispersion index \u003cstrong\u003eK: \u003c/strong\u003epercentage of chromosome with ratio=r\u0026gt;2;\u003c/p\u003e\n\u003cp\u003e\u003cstrong\u003eL:\u003c/strong\u003e ratio of largest to smallest chromosome; \u003cstrong\u003eM: \u003c/strong\u003edegree of asymmetry\u003c/p\u003e\n\u003cp\u003e\u003cstrong\u003e\u003cem\u003eC. diffusa\u003c/em\u003e\u003c/strong\u003e\u003cstrong\u003e from Jimma\u003c/strong\u003e\u003c/p\u003e\n\u003cp\u003eThe material of this species collected from Jimma possessed 2\u003cem\u003en\u003c/em\u003e=2\u003cem\u003ex\u003c/em\u003e= 30 (Table 1 and Figure 1G). According to centromeric ratio and chromosome length measurements, the karyotype (Figure 1H) has five metacentric pairs (pair numbers 1 to 5), 4 sub-metacentric pairs (pairs 6 to 9) and 6 sub-telocentric pairs (pair numbers 10 to 15) with a formula of 10m + 8sm + 12st. The asymmetry index is comparatively higher than the other studied \u003cem\u003eCommelina \u003c/em\u003especies with a value of 69.497 and according to Stebbins karyotype asymmetry classification, \u003cem\u003eC. diffusa \u003c/em\u003e(Jimma) belongs to 3A type (Table 1).\u003c/p\u003e\n\u003cp\u003eEven if no satellites were observed from the current plant material, a total of three interphase nucleoli were observed that possibly predict the number of NORs or satellites to be four (Figure 2F).\u003c/p\u003e\n\u003cp\u003e\u003cem\u003eC. diffusa \u003c/em\u003e(Jimma) possesses the smallest chromosomes size next to \u003cem\u003eC. subulata \u003c/em\u003ewith a length range from shortest 2.5\u0026micro;m to longest 4.58\u0026micro;m and total length of whole 2\u003cem\u003en \u003c/em\u003echromosome complement 104.22\u0026micro;m (Table 1). The total length of long arms for the diploid set was 72.43\u0026micro;m and total length of short arms for the diploid set was 31.79. The chromosomes range from very small to medium in size with an average length of 3.47\u0026micro;m and karyotype asymmetry of 2.28\u0026micro;m.\u003c/p\u003e\n\u003cp\u003eThis particular variant has the highest value in terms of intra-chromosomal asymmetry index (0.76717). It also possesses the lowest inter-chromosomal asymmetry index (0.13771), coefficient of variation in chromosome length (13.771), ratio in centromeric gradient (39.32) and dispersion index (5.415) when compared to values from all other studied plants of the genus \u003cem\u003eCommelina\u003c/em\u003e.\u003c/p\u003e\n\u003ch3\u003eC. subulata\u003c/h3\u003e\n\u003cp\u003eThe experimental species collected from Ginchi indicated that \u003cem\u003eC. subulata \u003c/em\u003ehas a diploid number of 2\u003cem\u003en \u003c/em\u003e=2\u003cem\u003ex\u003c/em\u003e= 30 (Table 1 and Figure 1I). As to arm ratio and chromosome length measurements, the karyotype (Figure 1J) consisted of nine metacentric pairs (pair numbers 1, 2, 3, 5, 8, 9, 10, 12, 13), five sub-metacentrics (4, 6, 11, 14, 15) and one pair of sub-telocentrics (7) with a karyotypic formula of 18m + 10sm + 2st. The species possesses the lowest value in asymmetry index (60.232), being the 2A type in Stebbins asymmetry classification (Table 1). Two interphase nucleoli were observed (Figure 2G), but no satellites were observed. One of the two nucleoli is larger than the other, which could result from fusion of two or more nucleoli; and it may be inferred that a total of four satellite chromosomes/ NORs are present.\u003c/p\u003e\n\u003cp\u003e\u003cem\u003eCommelina subulata \u003c/em\u003e(Ginchi) has the smallest chromosome size than other species included in this study with a length range from smallest1.923\u0026micro;m to largest 3.654\u0026micro;m and the total length of whole chromosome complement is 83.154\u0026micro;m (Table 1). .\u003c/p\u003e\n\u003cp\u003e\u0026nbsp;The total length of long arms and short arms for the diploid set was 50.085\u0026micro;m and 33.069\u0026micro;m, respectively (Table 1). The average karyotype asymmetry is 1.51 and the chromosomes, on the average arm ratio, can be considered as metacentric. The chromosomes are very small to medium in size with an average length of 2.77\u0026micro;m.\u003c/p\u003e\n\u003cp\u003e\u003cem\u003eC. subulata \u003c/em\u003epossesses the lowest intra-chromosomal asymmetry index, highest ratio of centromeric gradient and dispersion index with a value of 0.3523, 12.755 and 67.45, respectively. The species also consisted of an inter-chromosomal asymmetry index (0.1891) and coefficient of variation in chromosome length (18.91) (Table 1).\u003c/p\u003e"},{"header":"Discussion","content":"\u003cp\u003eIn the four species studied from the genus \u003cem\u003ecommelina\u003c/em\u003e, base numbers of \u003cem\u003ex\u003c/em\u003e=11 and \u003cem\u003ex\u003c/em\u003e=15 have been obtained. Base numbers \u003cem\u003ex\u003c/em\u003e=11, 13 and 15 was also previously reported for \u003cem\u003eCommelina \u003c/em\u003e[\u003ca href=\"#_ENREF_16\"\u003e16\u003c/a\u003e, \u003ca href=\"#_ENREF_17\"\u003e17\u003c/a\u003e].The species \u003cem\u003eCommelina benghalensis \u003c/em\u003e(2\u003cem\u003en\u003c/em\u003e=6\u003cem\u003ex\u003c/em\u003e=66) was based on \u003cem\u003ex\u003c/em\u003e=11 [\u003ca href=\"#_ENREF_18\"\u003e18\u003c/a\u003e]. Others, \u003cem\u003eC. africana\u003c/em\u003e, \u003cem\u003eC. diffusa \u003c/em\u003e(Jimma) and \u003cem\u003eC. subulata \u003c/em\u003ehaving chromosome number of (2\u003cem\u003en\u003c/em\u003e=2\u003cem\u003ex\u003c/em\u003e=30) were based on \u003cem\u003ex\u003c/em\u003e=15 base number. This result agrees with previous studies [\u003ca href=\"#_ENREF_15\"\u003e15\u003c/a\u003e, \u003ca href=\"#_ENREF_17\"\u003e17\u003c/a\u003e, \u003ca href=\"#_ENREF_19\"\u003e19-24\u003c/a\u003e].\u003c/p\u003e\n\u003cp\u003e\u003cem\u003eC. subulata \u003c/em\u003eof Ginchi has 2\u003cem\u003en\u003c/em\u003e=30 and this was in agreement with other reports from materials collected from Ethiopia [\u003ca href=\"#_ENREF_15\"\u003e15\u003c/a\u003e] and Ghana [\u003ca href=\"#_ENREF_25\"\u003e25\u003c/a\u003e]. Tetraploid species of \u003cem\u003eC. subulata \u003c/em\u003e2\u003cem\u003en\u003c/em\u003e=60 have also been reported for materials from India [\u003ca href=\"#_ENREF_26\"\u003e26\u003c/a\u003e, \u003ca href=\"#_ENREF_27\"\u003e27\u003c/a\u003e]. \u003cem\u003eC. diffusa \u003c/em\u003efrom Ginchi has 2\u003cem\u003en\u003c/em\u003e=66 and this number is reported for the first time.\u003c/p\u003e\n\u003cp\u003eChromosome numbers of \u003cem\u003eC. africana \u003c/em\u003ecollected from Addis Ababa and Sebeta were diploids 2\u003cem\u003en\u003c/em\u003e=30 and this was in agreement with previous reports [\u003ca href=\"#_ENREF_15\"\u003e15\u003c/a\u003e]. Even if, the current count was diploid chromosome number, Morton [\u003ca href=\"#_ENREF_28\"\u003e28\u003c/a\u003e] also found 2\u003cem\u003en\u003c/em\u003e=28 for materials from Ghana and polyploids (2\u003cem\u003en\u003c/em\u003e=60 and 2\u003cem\u003en\u003c/em\u003e= 120) were also reported [\u003ca href=\"#_ENREF_15\"\u003e15\u003c/a\u003e]. \u003cem\u003eC. benghalensis \u003c/em\u003eof Addis Ababa was found to be hexaploids (2\u003cem\u003en\u003c/em\u003e=6\u003cem\u003ex\u003c/em\u003e=66) and this was in agreement with earlier report [\u003ca href=\"#_ENREF_25\"\u003e25\u003c/a\u003e]. Both diploid 2\u003cem\u003en\u003c/em\u003e=22 and tetraploid (2\u003cem\u003en\u003c/em\u003e=44) cytotypes have also been reported for \u003cem\u003eC. benghalensis \u003c/em\u003efrom materials of Nigeria, China, India, Japan, Uganda, Ethiopia and Tanganyika [\u003ca href=\"#_ENREF_15\"\u003e15\u003c/a\u003e, \u003ca href=\"#_ENREF_18\"\u003e18-21\u003c/a\u003e, \u003ca href=\"#_ENREF_23\"\u003e23\u003c/a\u003e, \u003ca href=\"#_ENREF_25\"\u003e25\u003c/a\u003e, \u003ca href=\"#_ENREF_29\"\u003e29\u003c/a\u003e, \u003ca href=\"#_ENREF_30\"\u003e30\u003c/a\u003e]. A diploid \u003cem\u003eC. diffusa \u003c/em\u003e(2\u003cem\u003en\u003c/em\u003e=30) has been found for specimens collected from Jimma. This result was in agreement with other reports [\u003ca href=\"#_ENREF_15\"\u003e15\u003c/a\u003e, \u003ca href=\"#_ENREF_24\"\u003e24\u003c/a\u003e].\u003c/p\u003e\n\u003cp\u003ePolyploid series of the West African \u003cem\u003eC. benghalensis\u003c/em\u003e, \u003cem\u003eC. africana \u003c/em\u003eand \u003cem\u003eAneilema umbrsum \u003c/em\u003ecomplexes are of autopolyploid origin based on observation of close similarity between polyploid and diploids with the absence of allied taxa which could have been involved in allopolyploidy [\u003ca href=\"#_ENREF_25\"\u003e25\u003c/a\u003e].\u003c/p\u003e\n\u003cp\u003eA comparison made between the karyotypes of \u003cem\u003eC. diffusa \u003c/em\u003efrom Taiwan [\u003ca href=\"#_ENREF_23\"\u003e23\u003c/a\u003e] and \u003cem\u003eC. diffusa \u003c/em\u003e(Entoto mountain and Jimma) in the present study indicates that, although they share similar chromosome number (2\u003cem\u003en\u003c/em\u003e=30), they differ in karyotypic detail. The karyotype formula of the Taiwan specimen was reported with only two chromosomal groups, m and sm, (10m + 20sm), while in the present materials\u003c/p\u003e\n\u003cp\u003ethree chromosomal groups (m, sm and st) with a formula of 16m+6sm+8st (Entoto) and 10m + 8sm + 12st (Jimma) were observed. Alam and Sharma reported variation in karyotypes among five populations of Indian \u003cem\u003eC. diffusa \u003c/em\u003ehaving 2\u003cem\u003en \u003c/em\u003e= 30 [\u003ca href=\"#_ENREF_21\"\u003e21\u003c/a\u003e]. Four different karyotype formula within 2\u003cem\u003en\u003c/em\u003e=30 chromosomes from India were also reported previously [\u003ca href=\"#_ENREF_20\"\u003e20\u003c/a\u003e]. Factors, other than the chromosomal heteromorphism, like deviation in techniques of chromosome preparation, condensation difference and measurement technique can also lead to karyotype diversity among reports by different workers. These factors hinder comparison between various karyotypes and for real comparison mean values for each measurement of individual chromosomes must be taken. Furthermore, karyotype comparison for the remaining species was not performed as all the chromosome report matching to the current chromosome number had not been found.\u003c/p\u003e\n\u003cp\u003eThe chromosome report 2\u003cem\u003en\u003c/em\u003e=28 for the two species of \u003cem\u003eC. diffusa \u003c/em\u003eand \u003cem\u003eC. africana \u003c/em\u003ewas probably associated with the uncommon aneuploids formed due to loss of chromosome number [\u003ca href=\"#_ENREF_15\"\u003e15\u003c/a\u003e, \u003ca href=\"#_ENREF_28\"\u003e28\u003c/a\u003e]. But reduction in chromosome number can be associated with Robertsonian fusion [\u003ca href=\"#_ENREF_32\"\u003e32\u003c/a\u003e] and translocation of all or most of its part followed by loss of the chromosome. High rate of prevalence of aneuploidy and polyploidy in the genus \u003cem\u003eCommelina \u003c/em\u003ehad reported [\u003ca href=\"#_ENREF_25\"\u003e25\u003c/a\u003e].\u003c/p\u003e\n\u003cp\u003eJones and Joplings reported the chromosome size of the genus \u003cem\u003eCommelina \u003c/em\u003eas the smallest in the family [\u003ca href=\"#_ENREF_16\"\u003e16\u003c/a\u003e]. Faden and Morton, on the other hand, confirmed the presence of medium to relatively large chromosome within the genus [\u003ca href=\"#_ENREF_17\"\u003e17\u003c/a\u003e, \u003ca href=\"#_ENREF_25\"\u003e25\u003c/a\u003e]. In the present study, except for \u003cem\u003eC. subulata\u003c/em\u003e, the size was predominantly medium, whereas \u003cem\u003eC. subulata \u003c/em\u003ehas an average chromosome smaller than the medium size limit (3\u0026micro;m).\u003c/p\u003e\n\u003cp\u003eThe karyotype data of the present materials in the genus \u003cem\u003eCommelina \u003c/em\u003eindicated the presence of three types of chromosomes with higher frequency of metacentric (m) and sub-metacentric (sm) types than sub-telocentrics (st). This was supported by earlier reports [\u003ca href=\"#_ENREF_17\"\u003e17\u003c/a\u003e, \u003ca href=\"#_ENREF_25\"\u003e25\u003c/a\u003e]. However the ratio of each chromosome type considerably varies between each species.\u003c/p\u003e\n\u003cp\u003eA comparison made based on asymmetric index in the current study indicates that \u003cem\u003eC. africana \u003c/em\u003ecollected from Addis Ababa has same karyotype asymmetry with \u003cem\u003eC. diffusa \u003c/em\u003e(Jimma) which possess 65.19% and 65.56%, respectively. \u003cem\u003eC. benghalensis\u003c/em\u003e of the current study falls within 2B Stebbins category. This is in disagreement with the diploid material from Nigeria [\u003ca href=\"#_ENREF_30\"\u003e30\u003c/a\u003e]. This might be associated with genetic difference that associated with difference in agro-ecology.\u0026nbsp;\u003c/p\u003e\n\u003cp\u003eThe present study revealed some degree of variation in chromosome length between the studied species with no major chromosomal difference among themselves. This may be associated with differences in degree of chromosome condensation between the metaphase spread of different species measured. Thus, in practice, it is difficult to draw taxonomic conclusions simply by comparing chromosome length between taxa [\u003ca href=\"#_ENREF_33\"\u003e33\u003c/a\u003e] unless one compares chromosomes condensed to same degree.\u003c/p\u003e\n\u003cp\u003eSatellited chromosomes were observed frequently in Commelinaceae [\u003ca href=\"#_ENREF_25\"\u003e25\u003c/a\u003e]. In the present study, satellites were observed in \u003cem\u003eC. africana\u003c/em\u003e and \u003cem\u003eC. diffusa \u003c/em\u003e(Ginchi). The reason why satellites were not observed in some of the species may be that the satellites are too small and escape easy cytological detection, or condense. All the satellites detected were located at the tip of the short arm of chromosome. Discrepancies in number of satellites reported for a particular taxon or population can be due to the inability to observe all the satellites because of variation in techniques of chromosome preparation, stages at the time of chromosome analysis and chromosomal polymorphism [\u003ca href=\"#_ENREF_34\"\u003e34\u003c/a\u003e].\u003c/p\u003e\n\u003cp\u003eThe maximum number of telophase nucleoli obtained through silver staining may correspond to the maximum number of satellites that are present in particular taxon [\u003ca href=\"#_ENREF_34\"\u003e34\u003c/a\u003e]. Thus, even though all the satellites are not detected for various reasons discussed above, one can infer about the number of satellited chromosomes present in the taxon, if one is able to obtain the maximum number of nucleoli present. Accordingly it was assumed that the maximum number of satellites for \u003cem\u003eC. africana \u003c/em\u003e\u0026nbsp;will be four.\u003c/p\u003e\n\u003cp\u003eNucleoli are formed at NORs of the satellited chromosomes. During the cell cycle, nucleoli disappear at late prophase and reform during telophase. It is less easy to observe the maximum number of nucleoli during telophase. As the cell cycle proceeds from telophase to interphase, nucleoli tend to fuse together, and thus their number in most of the interphase nuclei is usually less than their number in telophase nuclei. Though it is less frequently, it is possible that the nucleus enters interphase without all the nucleoli being fused, in which case the maximum number of nucleoli can also be observed in interphase nuclei. The maximum number of nucleoli observed, be at telophase or interphase, can be used to infer about the number of active NORs the plant possesses. Even numbers of maximum nucleoli are expected because NORs chromosomes occur as homologous pair (s). In case the highest number observed is odd number, one may take the next higher even number as the number of nucleoli for the organism. Usually when odd number is observed, at least one of the nucleoli is larger than the rest of the nucleoli indicating that the large nucleolus is the product of fusion of smaller nucleoli. In the present study, maximum number of nucleoli observed for \u003cem\u003eC. africana \u003c/em\u003e(Addis Ababa) is three, and for \u003cem\u003eC. africana \u003c/em\u003e(sebeta) and \u003cem\u003eC. diffusa \u003c/em\u003e(Entoto) is four each which allow to make an inference that they all have 4 NORs (satellite chromosomes).\u003c/p\u003e\n\u003cp\u003eThe number of nucleoli observed for \u003cem\u003eC. benghalensis \u003c/em\u003eis five but two are very large relative to the other three nucleoli. The large ones are possibly fusion products of two or more small nucleoli. There are at least three pairs or more satellited chromosomes in this species.\u003c/p\u003e"},{"header":"Conclusions","content":"\u003cp\u003eThe present study has revealed and confirmed chromosome number, ploidy level, karyotype and nucleolus numbers of six species of Commelinaceae which were collected from different localities of central and south western part of the country. Accordingly, this study showed that basic chromosome number for three species of \u003cem\u003eCommelina \u003c/em\u003e(\u003cem\u003eC. diffusa, C. africana \u003c/em\u003eand \u003cem\u003eC. subulata\u003c/em\u003e) is \u003cem\u003ex\u003c/em\u003e=15 and 2\u003cem\u003en\u003c/em\u003e=30. The chromosomes of these species are predominantly of m and sm types. \u003cem\u003eC. benghalensis \u003c/em\u003e(2\u003cem\u003en\u003c/em\u003e=6\u003cem\u003ex\u003c/em\u003e=66) was also found to have \u003cem\u003ex\u003c/em\u003e=11. The basic chromosome number for \u003cem\u003eC. diffusa \u003c/em\u003e(Ginchi), 2\u003cem\u003en\u003c/em\u003e=66 was different from previous reports and hence it could be another cytotype for the species. Variation in karyotype formula is also observed within species of \u003cem\u003eC. africana \u003c/em\u003eand \u003cem\u003eC. diffusa \u003c/em\u003ecollected from different localities. \u003cem\u003eC. diffusa \u003c/em\u003e(Ginchi) is vigour, longer with distinct morphology than the other diploid species. This is the first work to present karyotypes, satellite chromosomes and nucleoli of the Ethiopian Commelinaceae. Nevertheless an integrated data from karyotype, molecular study of chloroplast genome and evolution through considering more representative species is necessary to infer phylogenetic relationship among the taxa.\u003c/p\u003e"},{"header":"Methods","content":"\u003cp\u003eFour species of the genus \u003cem\u003eCommelina \u003c/em\u003ewhich were collected from different localities of Central and Southwestern part of Ethiopia were analyzed cytologically. Chromosome study including numbers and morphology (karyotypes) was done on C-metaphase chromosomes of the root tip meristematic cells.\u003c/p\u003e\n\u003cp\u003e\u003cstrong\u003ePretreatment\u003c/strong\u003e\u003c/p\u003e\n\u003cp\u003eTo obtain clean roots for chromosome preparation, stems were harvested from potted plants and allowed to stand submerged in water. In a few days, roots emerged from the nodes of the submerged stems and harvested for metaphase arrest of mitosis. Hence, roots were immersed in 8-hydroxyquiniline (0.002M) for 3 to 5 hrs, fixed in 3:1 (v/v) of ethanol and glacial acetic acid 1-24 hr at about 4\u003csup\u003eo\u003c/sup\u003eC.\u003c/p\u003e\n\u003cp\u003e\u003cstrong\u003eMaceration\u003c/strong\u003e\u003c/p\u003e\n\u003cp\u003eRoots from fixative were rinsed several times in distilled water and macerated in a solution of pectinase and cellulase in a water bath at 37\u003csup\u003eo\u003c/sup\u003eC for about 1 hr or more. The process was stopped when the tips of the root started detaching from the root with or without agitation of the vial. Then, enzyme solution was carefully pipetted out and transferred to a watch glass or a petridish after rinsing.\u003c/p\u003e\n\u003cp\u003e\u003cstrong\u003eAir dry slide preparation\u003c/strong\u003e\u003c/p\u003e\n\u003cp\u003eOne or more of the macerated root tips (depending upon the size) were pipetted on a clean slide and fresh fixative (3:1, ethanol: acetic acid) were added to the root tips, and the tips were mashed quickly with flat ended mounted needle. The slide was then allowed to air dry at room temperature and stored until needed for staining.\u003c/p\u003e\n\u003cp\u003e\u003cstrong\u003eSlide staining\u003c/strong\u003e\u003c/p\u003e\n\u003cp\u003eSlides with good preparations were screened under phase contrast microscope and good preparations were screened under phase contrast microscope before staining. The promising preparation was stained in Giemsa stain in Sorenson phosphate buffer solution (PH 6.8). When correct contrast of staining was obtained, the slides were rinsed in distilled water, and allowed to air-dry at room temperature at least for 24 hrs. The preparation was made permanent by mounting in depex mountant and the gum was allowed to set for several days following by examining under camera fitted microscope. Then, enlarged photomicrograph prints were made accordingly.\u003c/p\u003e\n\u003cp\u003e\u003cstrong\u003eSilver staining\u003c/strong\u003e\u003c/p\u003e\n\u003cp\u003eSlide preparation was similar with the above with the exclusion of treatment of 8-hydroxyquiniline. The latter treatment was omitted in order to obtain telophase cells during which the maximum number of nucleoli may be observed. Silver staining technique was similar to the work of Dagne and Heneen [\u003ca href=\"#_ENREF_35\"\u003e35\u003c/a\u003e].\u003c/p\u003e\n\u003cp\u003e\u003cstrong\u003e\u0026nbsp;Karyotype analysis\u003c/strong\u003e\u003c/p\u003e\n\u003cp\u003eThe printed pictures of chromosomes were scanned into computer and the lengths of the whole chromosomes and their arms were measured in terms of pixel per cm using micro measure computer software version 3.3. Accordingly, the arm ratio of the chromosomes was calculated by dividing the length of the long arm to that of short arm.\u003c/p\u003e\n\u003cp\u003eKaryotypes were constructed by cutting and arranging the putative homologous chromosomes into pairs based on arm ratio(\u003cem\u003er\u003c/em\u003e) and chromosome size using the Smart Type software version 0.8. Chromosomes were categorized into chromosome types based on arm ratio (\u003cem\u003er\u003c/em\u003e) according to [\u003ca href=\"#_ENREF_36\"\u003e36\u003c/a\u003e] with slight modification. In the present case the term metacentric chromosome was used to include both M and m types with \u003cem\u003er \u003c/em\u003e=1.0-1.7. Sub-metacentric was used as synonymous with sub-median chromosome of \u003cem\u003er\u003c/em\u003e= 1.7-3.0 and st is similar to sub-telocentric when \u003cem\u003er\u003c/em\u003e=3.0-7.0.\u003c/p\u003e\n\u003cp\u003eAfter accurate measurements of karyotyped chromosomes were obtained, intra-chromosomal asymmetry (A\u003csub\u003e1\u003c/sub\u003e) and inter-chromosomal asymmetry (A\u003csub\u003e2\u003c/sub\u003e) indices have been calculated [\u003ca href=\"#_ENREF_37\"\u003e37\u003c/a\u003e]. Moreover, DI (dispersion index) of chromosomes was also calculated [\u003ca href=\"#_ENREF_38\"\u003e38\u003c/a\u003e].\u003c/p\u003e\n\u003cp\u003eMeasurements like intra chromosomal (A\u003csub\u003e1\u003c/sub\u003e), inter-chromosomal (A\u003csub\u003e2\u003c/sub\u003e), dispersion indices (DI) and Stebbins asymmetry depends on both chromosome size and centromeric position for estimating karyotype asymmetry of chromosomes. But asymmetry index depends only on centromeric position [\u003ca href=\"#_ENREF_39\"\u003e39\u003c/a\u003e].\u003c/p\u003e"},{"header":"Declarations","content":"\u003cp\u003e\u003cstrong\u003eEthics approval and consent to participate\u003c/strong\u003e\u003c/p\u003e\n\u003cp\u003eNot applicable\u003c/p\u003e\n\u003cp\u003e\u003cstrong\u003eConsent for publication\u003c/strong\u003e\u003c/p\u003e\n\u003cp\u003eNot applicable\u003c/p\u003e\n\u003cp\u003e\u003cstrong\u003eAvailability of data and materials\u003c/strong\u003e\u003c/p\u003e\n\u003cp\u003eThe data used for current study can be provided on request.\u003c/p\u003e\n\u003cp\u003e\u003cstrong\u003eCompeting interests\u003c/strong\u003e\u003c/p\u003e\n\u003cp\u003eThe authors declare that they have no competing interests\u003c/p\u003e\n\u003cp\u003e\u003cstrong\u003eFunding\u003c/strong\u003e\u003c/p\u003e\n\u003cp\u003eAll chemicals and materials required for the current study were funded from an internal grant of Aksum University.\u003c/p\u003e\n\u003cp\u003e\u003cstrong\u003eAuthors' contributions\u003c/strong\u003e\u003c/p\u003e\n\u003cp\u003eSG prepared the publication manuscript and all experimental works. GK, BG, GM and DG review all the work and incorporated necessary comments for better output.\u003c/p\u003e\n\u003cp\u003e\u003cstrong\u003eAcknowledgements\u003c/strong\u003e\u003c/p\u003e\n\u003cp\u003eIt is our great pleasure to acknowledge Dr. Kifle Dagne and Professor Ensermu Kelbessa of Addis Ababa University, for their meticulous assistance. We would also like to extend our heartfelt appreciation to Addis Ababa University, School of Graduate Studies, Faculty of Life Sciences for permission to access laboratory facilities to carry out our research.\u003c/p\u003e"},{"header":"References","content":"\u003col\u003e\n\u003cli\u003eKubitzki K, Rohwer J, Bittrich V: The families and genera of vascular plants, vol. 1: Springer; 1990.\u003c/li\u003e\n\u003cli\u003eKelbessa E, Demissew S: Diversity of vascular plant taxa of the flora of Ethiopia and Eritrea. \u003cem\u003eEthiopian Journal of Biological Sciences \u003c/em\u003e2014, 13(Supp.)):37-45.\u003c/li\u003e\n\u003cli\u003eFaden RB: New species of Commelina (Commelinaceae) from the Flora of Tropical East Africa. \u003cem\u003eNovon \u003c/em\u003e1994:224-235.\u003c/li\u003e\n\u003cli\u003eFaden RB: New or misunderstood species of Commelina (Commelinaceae) from the Flora of Tropical East Africa and Flora Zambesiaca areas. \u003cem\u003eNovon \u003c/em\u003e2001:398-409.\u003c/li\u003e\n\u003cli\u003eEdwards S, Demissew S, Hedberg I: Flora of Ethiopia and Eritrea: Hydrocharitaceae to Arecaceae: The National Herbarium, Addis Ababa University; 1997.\u003c/li\u003e\n\u003cli\u003eQureshi R, Bhatti GR: Ethnobotany of plants used by the Thari people of Nara Desert, Pakistan. \u003cem\u003eFitoterapia \u003c/em\u003e2008, 79(6):468-473.\u003c/li\u003e\n\u003cli\u003eVan der Burg W: Commelina benghalensis L. \u003cem\u003ePlant resources of tropical Africa \u003c/em\u003e2004, 2:212-214.\u003c/li\u003e\n\u003cli\u003eManandhar NP: Plants and people of Nepal: Timber press; 2002.\u003c/li\u003e\n\u003cli\u003eDeyuan H, Robert A: Commelina diffusa. \u003cem\u003eWu, ZY; Raven, PH; Hong, DY, Flora of China \u003c/em\u003e2000, 24.\u003c/li\u003e\n\u003cli\u003eFibrich B, Lall N: Commelina benghalensis. 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GENETICS SOC JAPAN NATL INST GENETICS, YATA, MISHIMA, SHIZUOKA 411, JAPAN: 340-\u0026amp;.\u003c/li\u003e\n\u003cli\u003eBhattacharya B: Cytological studies on some Indian members of Commelinaceae. \u003cem\u003eCytologia \u003c/em\u003e1975, 40(2):285-299.\u003c/li\u003e\n\u003cli\u003eAlam N: Trends of Chromosome evolution in family Commelinaceae. \u003cem\u003eThe Nucleus \u003c/em\u003e1984, 27:231-241.\u003c/li\u003e\n\u003cli\u003eEksomtramage L, Sirirugsa P, Sawangchote P, Jornead S, Saknimit T, Leeratiwong C: Chromosome number of some monocot species from Ton-Nga-Chang Wildlife Sanctuary, southern Thailand. \u003cem\u003eThai Forest Bulletin (Botany) \u003c/em\u003e2001(29):63-71.\u003c/li\u003e\n\u003cli\u003eFujishima H: Karyotypic diversity of Commelina benghalensis L.(Commelinaceae). \u003cem\u003eChromosome science \u003c/em\u003e2007, 10(2):43-53.\u003c/li\u003e\n\u003cli\u003eFujishima H: Karyotypic diversity of Commelina diffusa Burm.(Commelinaceae). \u003cem\u003eChromosome science \u003c/em\u003e2007, 10(1):21-28.\u003c/li\u003e\n\u003cli\u003eMorton JK: The Commelinaceae of West Africa: a biosystematic survey. \u003cem\u003eBot J Linn Soc \u003c/em\u003e1967, 60(382):167-221.\u003c/li\u003e\n\u003cli\u003eKammathy R, Rolla SR: Notes on Indian Commelinaceae-II: Cytological Observation. \u003cem\u003eNelumbo \u003c/em\u003e1961, 3(2):167-169.\u003c/li\u003e\n\u003cli\u003eRAGHAVAN RS: Cytological observation on the Indian species of Commelinaceae. \u003cem\u003eCurrent Sci \u003c/em\u003e1961, 30:310-311.\u003c/li\u003e\n\u003cli\u003eMorton J: Cytotaxonomic studies on the Gold Coast species of the genus Commelina Linn. \u003cem\u003eBot J Linn Soc \u003c/em\u003e1956, 55(361):507-531.\u003c/li\u003e\n\u003cli\u003eShigenobu Y, Kobori M: Karyotype variation of Commelina benghalensis correlated with habitats in Kochi Prefecture, Japan. \u003cem\u003eChromosome science \u003c/em\u003e1997, 1(2):83-87.\u003c/li\u003e\n\u003cli\u003eOziegbe M, Eludini PO: Karyotypic Studies of Commelina benghalensis variety benghalensis and C. forskalaei (Commelinaceae) from Nigeria. \u003cem\u003eCytologia \u003c/em\u003e2013, 78(2):151-156.\u003c/li\u003e\n\u003cli\u003ePanigraphi G: Cytotaxonomic studies in certain species of Commelina Linn. in eastern India. \u003cem\u003eJ Indian Bot Soc \u003c/em\u003e1964, 43:294-310.\u003c/li\u003e\n\u003cli\u003eGowda SR, Gawde HM, Hyderi A, Savitha MR, Krishnamurthy B, Karat SC, Doddaiah N, Patel ZM, Ramachandra NB: Chromosomal anomalies and congenital heart disease in Mysore, South India. \u003cem\u003eInt J Hum Genet \u003c/em\u003e2010, 10(1-3):131-139.\u003c/li\u003e\n\u003cli\u003eCarter S, Cutler D, Reynold T, Brandham P: A multidisciplinary approach to a revision of the Aloe somaliensis complex (Liliaceae). \u003cem\u003eKew bulletin \u003c/em\u003e1984:611-633.\u003c/li\u003e\n\u003cli\u003eDagne K: Karyotypes, C-banding and nucleolar numbers inGuizotia (Compositae). \u003cem\u003ePlant Syst Evol \u003c/em\u003e1995, 195(1-2):121-135.\u003c/li\u003e\n\u003cli\u003eDagne K, HENEEN WK: The karyotype and nucleoli of Guizotia abyssinica (Compositae). \u003cem\u003eHereditas \u003c/em\u003e1992, 117(1):73-83.\u003c/li\u003e\n\u003cli\u003eLevan A: Nomenclature for centromeric position on chromosomes. \u003cem\u003eHereditas \u003c/em\u003e1964, 52:201-220.\u003c/li\u003e\n\u003cli\u003eZarco CR: A new method for estimating karyotype asymmetry. \u003cem\u003eTaxon \u003c/em\u003e1986, 35(3):526-530.\u003c/li\u003e\n\u003cli\u003eLavania U, Srivastava S: Quantitative delineation of karyotype variation in Papaver as a measure of phylogenetic differentiation and origin. \u003cem\u003eCurr Sci \u003c/em\u003e1999:429-435.\u003c/li\u003e\n\u003cli\u003eGunjan K, Roy BK: Karyotype studies in dominant species of Aloe from eastern India. \u003cem\u003eCaryologia \u003c/em\u003e2010, 63(1):41-49.\u003c/li\u003e\n\u003c/ol\u003e"}],"fulltextSource":"","fullText":"","funders":[],"hasAdminPriorityOnWorkflow":false,"hasManuscriptDocX":true,"hasOptedInToPreprint":true,"hasPassedJournalQc":"","hasAnyPriority":false,"hideJournal":true,"highlight":"","institution":"","isAcceptedByJournal":false,"isAuthorSuppliedPdf":false,"isDeskRejected":"","isHiddenFromSearch":false,"isInQc":false,"isInWorkflow":false,"isPdf":false,"isPdfUpToDate":true,"isWithdrawnOrRetracted":false,"journal":{"display":true,"email":"[email protected]","identity":"researchsquare","isNatureJournal":false,"hasQc":true,"allowDirectSubmit":true,"externalIdentity":"","sideBox":"","snPcode":"","submissionUrl":"/submission","title":"Research Square","twitterHandle":"researchsquare","acdcEnabled":true,"dfaEnabled":false,"editorialSystem":"","reportingPortfolio":"","inReviewEnabled":false,"inReviewRevisionsEnabled":true},"keywords":"Commelina L., Ethiopia, Karyotype, Nucleoli, Satellite","lastPublishedDoi":"10.21203/rs.3.rs-100645/v1","lastPublishedDoiUrl":"https://doi.org/10.21203/rs.3.rs-100645/v1","license":{"name":"CC BY 4.0","url":"https://creativecommons.org/licenses/by/4.0/"},"manuscriptAbstract":"\u003cp\u003eBackground\u003c/p\u003e\u003cp\u003eWith about 100 species, \u003cem\u003eCommelina \u003c/em\u003eis the largest genus of Commelinaceae in Africa. Although medicinal and economic benefits had been studied extensively, little is known about its cytological analysis. Hence, this study will focus on chromosome and nucleoli analysis of selected species of Commelina L. Somatic chromosomes were prepared from root tips that emerged from the nodes of stem cuttings that were made to stand submerged in water. The roots were pretreated in 8-hydroxyquinelin 3-5 hrs followed by fixation in 3:1 ethanol: acetic acid for 1-24 hrs at 4\u003csup\u003eO\u003c/sup\u003eC. Air-dry slides were prepared following cellulase and pectinase maceration at 37\u003csup\u003eO\u003c/sup\u003eC, the preparation was stained in Giemsa stain (PH 6.4), rinsed and mounted. Nucleoli were stained in silver nitrate solution. \u003c/p\u003e\u003cp\u003eResults\u003c/p\u003e\u003cp\u003eChromosome numbers and Karyotype formula of the four species were found as \u003cem\u003eC. africana \u003c/em\u003e2\u003cem\u003en\u003c/em\u003e=2\u003cem\u003ex\u003c/em\u003e=30 (12m + 10sm + 8st), \u003cem\u003eC. benghalensis \u003c/em\u003e2\u003cem\u003en\u003c/em\u003e=6\u003cem\u003ex\u003c/em\u003e=66 (36m + 24sm + 6st), \u003cem\u003eC. diffusa \u003c/em\u003e(Ginchi) 2\u003cem\u003en\u003c/em\u003e=66 (28m +26sm + 12st), \u003cem\u003eC. diffusa (\u003c/em\u003eJimma\u003cem\u003e) \u003c/em\u003e2\u003cem\u003en\u003c/em\u003e=2\u003cem\u003ex\u003c/em\u003e=30 (10m + 8sm + 12st) and \u003cem\u003eC. subulata \u003c/em\u003e2\u003cem\u003en\u003c/em\u003e=2\u003cem\u003ex\u003c/em\u003e=30 (18m +10sm + 2st). According to Stebbins karyotype asymmetry, the karyotypes of \u003cem\u003eC. africana\u003c/em\u003e and \u003cem\u003eC. subulata \u003c/em\u003ewere 2A type, while that of \u003cem\u003eC. benghalensis \u003c/em\u003eand \u003cem\u003eC. diffusa \u003c/em\u003e(Ginchi) were 2B type. 3A asymmetry type was obtained for \u003cem\u003eC. diffusa \u003c/em\u003e(Jimma). Karotypes of \u003cem\u003eTradescantia \u003c/em\u003ewere found to be monomodal for the \u003cem\u003eCommelina \u003c/em\u003especies of the studied plant materials. Satellites were observed for species \u003cem\u003eC. africana\u003c/em\u003e and \u003cem\u003eC. diffusa \u003c/em\u003ewith variation in number ranging from 2 to 6. The maximum number of nucleoli observed varies from two to four for \u003cem\u003eCommelina\u003c/em\u003e. \u003c/p\u003e\u003cp\u003eConclusions\u003c/p\u003e\u003cp\u003e\u003cspan class=\"ql-cursor\"\u003e\u003c/span\u003eThis study reported karyotype and nucleoli of the Ethiopian Commelinaceae for the first time. The current investigation can be considered as an additional karyotype data to the earlier meiosis report for Ethiopian materials.\u003c/p\u003e","manuscriptTitle":"Karyo-Morphology and Nucleoli Analysis of Commelina L. (Commelinaceae) from Ethiopia","msid":"","msnumber":"","nonDraftVersions":[{"code":1,"date":"2020-11-04 15:18:04","doi":"10.21203/rs.3.rs-100645/v1","editorialEvents":[{"type":"communityComments","content":0}],"status":"published","journal":{"display":true,"email":"[email protected]","identity":"researchsquare","isNatureJournal":false,"hasQc":true,"allowDirectSubmit":true,"externalIdentity":"","sideBox":"","snPcode":"","submissionUrl":"/submission","title":"Research Square","twitterHandle":"researchsquare","acdcEnabled":true,"dfaEnabled":false,"editorialSystem":"","reportingPortfolio":"","inReviewEnabled":false,"inReviewRevisionsEnabled":true}}],"origin":"","ownerIdentity":"8e8f50e9-4e85-4af2-ac84-9781d956e435","owner":[],"postedDate":"November 4th, 2020","published":true,"recentEditorialEvents":[],"rejectedJournal":[],"revision":"","amendment":"","status":"posted","subjectAreas":[{"id":975281,"name":"Molecular Genetics"}],"tags":[],"updatedAt":"2020-11-04T15:18:05+00:00","versionOfRecord":[],"versionCreatedAt":"2020-11-04 15:18:04","video":"","vorDoi":"","vorDoiUrl":"","workflowStages":[]},"version":"v1","identity":"rs-100645","journalConfig":"researchsquare"},"__N_SSP":true},"page":"/article/[identity]/[[...version]]","query":{"redirect":"/article/rs-100645","identity":"rs-100645","version":["v1"]},"buildId":"7rjqhiLT3MXkJMwkYKINL","isFallback":false,"isExperimentalCompile":false,"dynamicIds":[84888],"gssp":true,"scriptLoader":[]}