Carnosic acid: A potent chemopreventive agent against oral carcinogenesis

The present study was aimed to investigate the chemopreventive potential of carnosic acid in 7,12-dimethylbenz(a)anthracene (DMBA)-induced hamster buccal pouch carcinogenesis. The chemopreventive potential was assessed by analyzing the tumor incidence, tumor volume and burden as well as by measuring the status of lipid peroxidation, non-enzymatic and enzymatic antioxidants and phase I and phase II detoxification enzymes. Oral squamous cell carcinoma was developed in the buccal pouch of golden Syrian hamsters by painting with 0.5% DMBA in liquid paraffin three times a week for 14 weeks. In the present study, 100% tumor formation was observed in hamsters treated with DMBA alone. Also, the status of lipid peroxidation, antioxidants and phase I and phase II detoxification enzymes were significantly altered during DMBA-induced oral carcinogenesis. Oral administration of carnosic acid at a dose of 10 mg/kg body weight/day to DMBA-treated animals completely prevented the tumor formation in the hamsters’ buccal pouches. Also, carnosic acid exerted potent anti-lipid peroxidative function and stimulated the detoxification cascade during DMBA-induced hamster buccal pouch carcinogenesis. The results of the present study suggest that the chemopreventive potential of carnosic acid is probably due to its anti-lipid peroxidative potential and modulating effect on carcinogen detoxification enzymes during DMBA-induced oral carcinogenesis.     

Evaluation of chemopreventive effects of Thymoquinone on cell surface glycoconjugates and cytokeratin expression during DMBA induced hamster buccal pouch carcinogenesis

The present study aimed to investigate the membrane stabilizing effect of Thymoquinone (TQ) on cell surface glycoconjugates and cytokeratin expression against DMBA induced hamster buccal pouch carcinogenesis. 0.5% DMBA painting (three times per week) in hamster buccal pouches for 14 weeks resulted in the formation of well developed oral squamous cell carcinoma. We observed 100% tumor formation with marked abnormalities of glycoconjugates status in tumor bearing hamsters as compared to control animals. Oral administration of TQ at a dose of 30 mg/kg body weight, to DMBA painted hamsters on alternate days for 14 weeks, reduced the tumor formation as well as protected the levels of cell surface glycoconjugates in DMBA painted hamsters. The present study thus suggests that TQ has potent chemopreventive efficacy as well as protected the abnormalities on cell surface glycoconjugates during DMBA induced hamster buccal pouch carcinogenesis.     

Chemopreventive efficacy of geraniol against 7,12-dimethylbenz[a]anthracene-induced hamster buccal pouch carcinogenesis

Abstract

The status of lipid peroxidation, antioxidants, and detoxification enzymes were used as biochemical end points to assess the chemopreventive potential of geraniol, a monoterpene, in 7,12-dimethylbenz[a]anthracene (DMBA)-induced hamster buccal pouch carcinogenesis. Topical application of 0.5% DMBA in liquid paraffin, three times a week, for 14 weeks developed well-differentiated squamous cell carcinoma in the buccal pouch of golden Syrian hamsters. Although 100% tumor formation was noticed in hamsters treated with DMBA alone, intragastric administration of geraniol, at a dose of 250 mg/kg body weight (b.w.) to DMBA-treated hamster completely prevented the formation of oral tumors. Furthermore, geraniol significantly reduced lipid peroxidation by-products and improved the status of enzymatic and non-enzymatic antioxidants as well as modulated the status of phase I and phase II detoxification enzymes, favoring the excretion of carcinogenic metabolite, during DMBA-induced oral carcinogenesis. The present study concludes that the chemopreventive potential of geraniol relies on its anti-lipid peroxidative and antioxidant function as well as modulatory effects on phase I and II detoxification enzymes to excrete the carcinogenic metabolite, during DMBA-induced hamster buccal pouch carcinogenesis.     

Chemopreventive efficacy of geraniol against 7,12-dimethylbenz[a]anthracene-induced hamster buccal pouch carcinogenesis

The status of lipid peroxidation, antioxidants, and detoxification enzymes were used as biochemical end points to assess the chemopreventive potential of geraniol, a monoterpene, in 7,12-dimethylbenz[a]anthracene (DMBA)-induced hamster buccal pouch carcinogenesis. Topical application of 0.5% DMBA in liquid paraffin, three times a week, for 14 weeks developed well-differentiated squamous cell carcinoma in the buccal pouch of golden Syrian hamsters. Although 100% tumor formation was noticed in hamsters treated with DMBA alone, intragastric administration of geraniol, at a dose of 250 mg/kg body weight (b.w.) to DMBA-treated hamster completely prevented the formation of oral tumors. Furthermore, geraniol significantly reduced lipid peroxidation by-products and improved the status of enzymatic and non-enzymatic antioxidants as well as modulated the status of phase I and phase II detoxification enzymes, favoring the excretion of carcinogenic metabolite, during DMBA-induced oral carcinogenesis. The present study concludes that the chemopreventive potential of geraniol relies on its anti-lipid peroxidative and antioxidant function as well as modulatory effects on phase I and II detoxification enzymes to excrete the carcinogenic metabolite, during DMBA-induced hamster buccal pouch carcinogenesis.     

Influence of beta-carotene on lysosomal hydrolases and their natural substrates in major salivary glands of hamsters treated with 7,12-dimethylbenzanthracene (DMBA)

We evaluated the effects of beta-carotene, a precursor of vitamin A, on the activity of some lysosomal hydrolases and on the levels of their natural substrates in hamster major salivary glands during experimental oral 7,12-dimethylbenzanthracene (DMBA) carcinogenesis. Sixty-four hamsters (Cricetus auratus) were divided into four groups--group 1: untreated control; group 2: DMBA was painted three times a week in the left buccal pouch; group 3: beta-carotene was painted three times a week in the left buccal pouch; group 4: DMBA and beta-carotene were painted alternatively in the left buccal pouch. After 16 weeks, the animals were sacrificed and the activities of some lysosomal hydrolases and their natural substrates in the major salivary glands were measured. beta-Carotene when administered topically in DMBA treated animals (group 4) reduced the levels of the majority of enzymes and substrates closer to those of the untreated control group, thus outlining a mild protective effect of beta-carotene towards the DMBA carcinogenic stress. Nevertheless, the presence of some enzymes which responded negatively to the combined administration of DMBA and beta-carotene suggests the necessity for future studies on the effect of beta-carotene at different concentrations, the systemic administration and the possibility to combine the topical beta-carotene administration with other chemopreventive drugs.     

Ethanolic leaf extract of neem (Azadirachta indica) inhibits buccal pouch carcinogenesis in hamsters

We evaluated the chemopreventive effects of ethanolic neem leaf extract in the initiation and post-initiation phases of 7,12-dimethylbenz[a]anthracene (DMBA)-induced hamster buccal pouch (HBP) carcinogenesis. The frequency of bone marrow micronuclei as well as the concentrations of lipid peroxides, ratio of reduced to oxidized glutathione (GSH/GSSG), and the activities of the GSH-dependent enzymes glutathione peroxidase (GPx) and glutathione-S-transferase (GST) in the buccal pouch, liver and erythrocytes were used as biomarkers of chemoprevention. All the hamsters painted with DMBA alone for 14 weeks developed buccal pouch carcinomas that showed diminished lipid peroxidation and enhanced antioxidant status associated with increased frequencies of bone marrow micronuclei. In the liver and erythrocytes of tumour-bearing animals, enhanced lipid peroxidation was accompanied by compromised antioxidant defences. Administration of ethanolic neem leaf extract effectively suppressed DMBA-induced HBP carcinogenesis as revealed by the absence of tumours in the initiation phase and reduced tumour incidence in the post-initiation phase. In addition, ethanolic neem leaf extract modulated lipid peroxidation and enhanced antioxidant status in the pouch, liver and erythrocytes and reduced the incidence of bone marrow micronuclei. The results of the present study, demonstrate that ethanolic neem leaf extract inhibits the development of DMBA-induced HBP tumours by protecting against oxidative stress.     

Effect of curcumin and ferulic acid on modulation of expression pattern of p53 and bcl-2 proteins in 7,12-dimethylbenz[a]anthracene-induced hamster buccal pouch carcinogenesis

The modulating effect of curcumin and ferulic acid was investigated on expression pattern of apoptosis regulatory p53 and bcl-2 proteins in oral squamous cell carcinoma (OSCC). The OSCC was induced in the buccal pouch of golden Syrian hamster by painting with 0.5% 7,12-dimethylbenz[a]anthracene (DMBA) three-times a week for 14 weeks. The expression pattern of p53 and bcl-2 proteins was analyzed by immunohistochemical staining. We noticed 100% tumor formation in hamsters painted with DMBA alone for 14 weeks. Overexpression of p53 and bcl-2 proteins was observed in the buccal mucosa of tumor-bearing hamsters. Oral administration of curcumin (80 mg/kg body wt) and ferulic acid (40 mg/kg body wt) to DMBA painted hamsters on days alternate to DMBA painting for 14 weeks completely inhibited tumor formation and down-regulated the expression pattern of p53 and bcl-2 proteins. Our results thus demonstrated the protective role of curcumin and ferulic acid on DMBA-induced abnormal expression of p53 and bcl-2 proteins in the buccal mucosa of golden Syrian hamsters.     

Paeonol exhibits anti-tumor effects by apoptotic and anti-inflammatory activities in 7,12-dimethylbenz(a)anthracene induced oral carcinogenesis

We investigated the preventive potential of paeonol on 7,12-dimethylbenz(a)anthracene (DMBA) induced oral carcinogenesis. Oral tumors were developed in the buccal pouches of Syrian golden hamsters using topical application of 0.5% DMBA three times/week for 10 weeks. DMBA treated hamsters developed hyperplasia, dysplasia and well-differentiated squamous cell carcinoma. The animals also exhibited increased lipid oxidation, decreased antioxidant status and altered levels of detoxification agents. Paeonol treatment of DMBA treated hamsters for 14 weeks decreased tumor incidence, volume and burden Paeonol treatment also increased antioxidant activity and decreased lipid oxidation to near normal levels. Histomorphology and the expression patterns of mutant p53, cyclo-oxygenase (COX-2) and caspase-9 were investigated in the oral buccal mucosa. Paeonol exhibited protective effects against DMBA induced oral carcinogenesis owing to its antitumor, antioxidant, anti-inflammatory and apoptosis inducing properties.     

Antiproliferative and apoptosis inducing effect of lactoferrin and black tea polyphenol combination on hamster buccal pouch carcinogenesis

Combination chemoprevention using tea polyphenols as one of the components has received growing consideration in recent years. The present study was designed to evaluate the antiproliferative and apoptosis inducing effects of bovine lactoferrin (bLF) and black tea polyphenol (Polyphenon-B: P-B) combination on 7,12-dimethylbenz[a]anthracene (DMBA)-induced hamster buccal pouch (HBP) carcinogenesis. Topical application of DMBA for 14 weeks induced buccal pouch tumours that showed aberrant expression of cytokeratins, a marker for epithelial carcinomas. This was associated with increased cell proliferation and evasion of apoptosis as revealed by upregulation of proliferating cell nuclear antigen, NF-kappaB, mutant p53, Bcl-2 and downregulation of Bax, Fas and caspase 3 protein expression. Although dietary administration of bLF and Polyphenon-B alone significantly reduced tumour incidence, combined administration of bLF and Polyphenon-B was more effective in inhibiting HBP carcinogenesis by restoring normal cytokeratin expression, inhibiting cell proliferation and inducing apoptosis. These findings suggest that a "designer item" approach will be useful for human oral cancer prevention strategies.     

Beta carotene is associated with the regression of hamster buccal pouch carcinoma and the induction of tumor necrosis factor in macrophages

Beta carotene (250 micrograms/ml) dissolved in mineral oil applied either topically or injected locally (190 ng/ml dissolved in media) into DMBA (7,12-dimethylbenz(a)anthracene)-induced or HCPC-1 cell line-produced oral squamous cell carcinoma of the hamster buccal pouch was observed to result in the regression of these tumors. (p less than or equal to .005) Beta carotene application to tumor bearing pouches was observed to produce a dramatic increase in positively stained macrophages for tumor necrosis factor (TNF-alpha) as compared to macrophages in control pouches. Macrophages from hamsters with regressed tumor were shown to produce a significant increase in cytotoxicity to HCPC-1 tumor cells. Regression of the hamster oral carcinoma was correlated with the increased capacity of macrophages to lyse tumor cells, and related to the induction of tumor necrosis factor which was associated with the administration of the carotenoid, beta carotene.     

The inhibitory effect of oral administration of garlic on experimental carcinogenesis in hamster buccal pouches by DMBA painting

The inhibitory effect of oral administration of garlic on experimental carcinogenesis in buccal pouches induced by painting 9,10-dimethyl-1,2-benz(a)anthracene (DMBA) was studied on 40 golden Syrian hamsters. The animals were grouped at random into four experimental groups (oral administration of garlic, NTP, BP or mineral oil followed by DMBA painting on buccal pouches), three chemical control groups (oral administration of garlic, NTP or BP without DMBA painting) and a DMBA control group (only painted DMBA on buccal pouches). Starting from the fourth week after DMBA painting, the pouch mucosae were examined biweekly for its tumor formation and blood vessel architecture. Animals were sacrificed 25 weeks after DMBA application. Tumors and pouch mucosae were dissected to examine tumor nature and biochemical reactions of DNA synthesis and GGTase activity. The inhibitory efficacy of garlic, BP and NTP were evaluated according to the results of these examinations. Garlic was found to have a higher inhibitory efficacy than BP and NTP through the probable mechanism of competitive binding with nuclear DNA and diminishing the opportunity of DMBA to initiate carcinogenesis. Other factors related to cancer inhibition included insufficient local blood flow, low GGTase activity and lesser DNA synthesis. The inhibitory effect of fractions of garlic on experimental carcinogenesis should be a reasonable and necessary continuation in future studies of the series of cancer prevention by garlic.     

Apoptosis induction by S-allylcysteine,a garlic constituent,during 7,12-dimethylbenz[a]anthracene-induced hamster buccal pouch carcinogenesis

The apoptosis-inducing capacity of S-allylcysteine (SAC), a water-soluble garlic constituent, during 7,12-dimethylbenz[a]anthracene-induced hamster buccal pouch (HBP) carcinogenesis was investigated in male Syrian hamsters using DNA fragmentation and the apoptosis-associated proteins, tissue transglutaminase (tTG) and Bcl-2. Hamsters were divided into four groups of six animals each. Animals in group 1 were painted with a 0.5% solution of DMBA in liquid paraffin on the right buccal pouches three times a week for 14 weeks. Group 2 animals painted with DMBA as in group 1, in addition received 200 mg kg(-1) body weight SAC orally on days alternate to DMBA application. Group 3 animals received SAC as in group 2. Group 4 animals received neither DMBA nor SAC and served as the control. The experiment was terminated at the end of 14 weeks. Administration of SAC (200 mg kg(-1) body weight) to animals painted with DMBA inhibited DMBA-induced HBP carcinogenesis as revealed by the absence of neoplasms, induction of tTG and inhibition of Bcl-2 expression. The results of the present study suggest that SAC may exert its chemopreventive effect by inducing apoptosis.     

Establishment of two cell lines from hamster buccal pouch tumors induced by topical 7,12-dimethylbenz(a)anthracene and topical DMBA in conjunction with herpes simplex virus infection

Summary Two cell lines designated HBPC-1 and HBPC-2 have been established from hamster buccal pouch tumors induced by topical 7,12-dimethylbenz(a)anthracene (DMBA) and DMBA in conjunction with type 1 herpes simplex virus infection, respectively. The cells are epithelial in morphology, have a doubling time of approximately 18 h, and require bovine serum for optimal growth. The karyotype is aneuploid, with several marker chromosomes, and the cells produce squamous cell carcinomas when transplanted into normal hamster pouch tissues. The study is partly supported by grants from the Smokeless Tobacco Research Council, Inc., #0052 and 0061, and from NIDR #007323.     

Topical Treatment of Dermatophytic Lesion on Mice (<Emphasis Type="Italic">Mus musculus</Emphasis>) Model

Antidermatophytic potential of three weed plants viz. Tridax procumbens L., Capparis decidua (forsk) Edgew and Lantana camara L. were explored and experimentally induced dermatophytic lesion was topically treated in mice. Microbroth dilution method was carried out for determination of MIC and MFC of different extracts of selected plants. In animal studies, mice were experimentally inoculated with Trichophyton mentagrophytes and infected animals were topically treated with 5 mg/g terbinafine and two concentrations, i.e., 5 and 10 mg/g of test extract ointment. Complete recovery from the infection was observed on 12th day of treatment for reference drug terbinafine (5 mg/g) and 10 mg/g concentration of test extract ointment whereas 5 mg/g concentration of test extract ointment showed complete cure on 16th day of treatment. Fungal burden was also calculated by culturing skin scrapings from infected animals of different groups. Test extract ointment successfully treated induced dermatophytosis in mice without any disease recurrence incidences, thereby indicating efficacy of test extract as an excellent topical antifungal agent for the cure of dermatophytosis.     

Establishment and characterization of a cell line (HCDB-1) derived from a hamster buccal pouch carcinoma induced by DMBA and Taiwanese betel quid extract

This study identified that the carcinogenesis of hamster buccal pouch (HBP) induced by 7,12-dimethylbenz[a]anthracene (DMBA) was greatly enhanced (18 folds) by a combination treatment with Taiwanese betel quid (BQ) extract. A new cell line, HCDB-1, has been established from induced carcinomas. The cultured monolayer cells were epithelioid in shape with irregular nuclei. They demonstrated abundant cytokeratin and tonofilaments; however, ultrastructural well-organized desmosomes were lacking. The HCDB-1 cell exhibited population doubling in 19 h and was highly tumorigenic in nude mice. A C-->T transition at codon 141 (Ala to Val) of the p53 gene was detected in this cell. This mutation is equivalent to a specific temperature-sensitive mouse p53Ala135Val mutant that causes transformation by shifting to 37.5 degrees C. HCDB-1 is the first cell line established from the HBP model of oral carcinogenesis induced by DMBA/Taiwanese BQ extract. It might be valuable for exploring the molecular pathogenesis of oral cancer.     

广西三种真红树植物可培养细菌多样性及其生物活性初筛

为了挖掘真红树植物潜在细菌新物种和生物活性物质,丰富红树林微生物多样性,为新型活性产物开发提供菌株资源。该文从秋茄、木榄和红海榄三种广西来源的真红树植物及其生境中,按根、茎、叶、花、果实和泥土分成22份样品,选用8种不同培养基分离可培养细菌,通过16S rRNA基因序列鉴定,分析其多样性,采用纸片法筛选细菌发酵粗提物的抑菌活性,点植法测试其酶活性。结果表明：(1)共分离获得可培养细菌35株,隶属于23个科28个属,芽孢杆菌属占细菌总数的14.3%,为优势菌属,同时发现11株潜在的新细菌资源。(2)活性筛选获得4株细菌具有抑菌活性,16株细菌具有酶活性,芽孢杆菌属是酶活性优势菌属。综上所述,广西真红树植物可培养细菌多样性丰富,部分细菌具有抑菌活性和酶活性,在新型抗生素和酶应用方面具有一定的开发潜力。     

Suppression of tumor growth and invasion in 9,10 dimethyl benz(a) anthracene induced mammary carcinoma by the plant bioflavonoid quercetin

Administration of quercetin, a common polyphenolic component of many vascular and edible plants including vegetables, fruits and tea significantly reduced the tumor volume in rats induced for mammary carcinoma using dimethyl benz (a) anthracene (DMBA). Dose response was assessed, by treating the animals with different doses (15-45 mg/kgbw) of quercetin and 25 mg/kgbw was taken as effective dose. Quercetin was administered as an intra tumoral injection once a week for 4 weeks. Serum levels of carcino embryonic antigen (CEA), a potent marker for tumor growth and invasion was significantly decreased on quercetin treatment. Quercetin caused a significant decrease in the activities of acid phosphatase and Cathepsin D in serum of experimental animals. Activities of lysosomal enzymes- (beta-D galactosidase, beta-D glucuronidase, beta-D glucosidase and sialidase), in serum and tissue were significantly altered in DMBA animals compared to control animals. However, quercetin treatment caused no significant change in lysosomal enzyme activities in tissues, whereas the activities were significantly lowered in serum. Partial purification of tissue type plasminogen activator (t-PA) from the tumor and kidney showed increased activity in the DMBA induced animals. Serum urokinase, -like plasminogen activator (u-PA) was also increased in animals with tumor, indicating tumor invasion. Administration of quercetin caused a significant decrease of both t-PA and u-PA. In conclusion, the present study suggests the possible role of quercetin in primary and invasive mammary tumor treatment. The above observations in vivo warrant further studies, due to the easy availability, common occurrence and low toxicity of this dietary bioflavonoid.     

Effects of aqueous cinnamon extract on chemically-induced carcinoma of hamster cheek pouch mucosa

This study aimed to investigate the effects of aqueous cinnamon extract (ACE) on 7, 12-Dimethylbenz[a]anthracene (DMBA)-induced oral carcinogenesis in hamster cheek pouch (HCP) mucosa. Sixty male Syrian hamsters were randomly divided into six equal groups. The hamsters of groups I, II and III received no treatment, DMBA and ACE respectively, for 16 weeks. Groups IV and V were handled as group II and concomitantly treated with ACE for the same period and additionally group V received ACE for other 16 weeks after the stoppage of DMBA application. Group VI hamsters were handled as group III and additionally received DMBA for other 16 weeks after the stoppage of ACE supplementation. Hamsters of each group were euthanized according to the experimental schedule. The buccal pouches were and prepared for H&E stain, PAS reagent, CD3 and PDGF immunohistochemical reactivity. All groups showed dysplastic changes with varying degrees except groups I and III. Deep invasive carcinomas were recorded in 90% of the samples of group II, 60% of group IV, 50% of group V and 40% of group VI. From the previous results, it can be concluded that ACE has the potentiality preventing oral cancer initiation better than inhibiting oral cancer progression.     

Low level X-radiation effects on carcinogenesis by 7,12-dimethylbenz(a)anthracene in Syrian hamster cheek pouch epithelium: acute vs fractionated radiation dose studies

Studies examined the effects of acute and fractionated low to moderate level X-ray exposures on hamster cheek pouch carcinogenesis in vivo by 7,12-dimethylbenz(a)anthracene (DMBA). Animals were grouped by treatment as follows: acute doses of 0.85-3.40 Gy X rays; 17 once weekly doses of 0.01-0.20 Gy X rays (fractionated radiation); topical DMBA for 10 weeks; DMBA plus fractionated radiation starting together; DMBA plus acute radiation in Week 1 or 10 of DMBA treatments; and sham irradiation, DMBA vehicle, or anesthesia controls. After 44 weeks, hamsters were sacrificed, and their cheek pouches were excised, serially sectioned, and examined by light microscopy for histopathology. No histologic changes were observed in radiation-only hamsters. Carcinoma incidences in DMBA-only groups ranged from 45 to 60%. Carcinoma incidences were greater in groups receiving DMBA plus fractionated radiation than in groups receiving either acute radiation + DMBA or DMBA alone. Carcinoma incidences in acute radiation plus DMBA groups were lower than those in DMBA-only groups. These results suggest complex interactions between radiation and DMBA, perhaps with radiogenic cell killing being a principal factor in acute radiation + DMBA groups, and reciprocal additive or synergistic effects of radiation and DMBA on cancer induction and manifestation in fractionated radiation + DMBA groups.     

Population genetic structure of three tree species in the mangrove genus Ceriops (Rhizophoraceae) from the Indo West Pacific

Ceriops is a viviparous mangrove with widespread species Ceriops decandra and C. tagal, and an endemic species C. australis. Genetic diversity of the three species was screened in 30 populations collected from 23 locations in the Indo West Pacific (IWP) using Inter-simple sequence repeats (ISSR) and sequences of partial nuclear gene (G3pdh) and chloroplast DNA (trnV-trnM). At the species level, the total gene diversity (Ht) revealed by ISSRs was 0.270, 0.118, and 0.089 in C. decandra, C. tagal, and C. australis, respectively. A total of six haplotypes of G3pdh and five haplotypes of trnV-trnM were recognized among the three species. Only C. decandra was detected containing more than one haplotype from each sequence data set (four G3pdh haplotypes and three trnV-trnM haplotypes). At the population level, genetic diversity of Ceriops was relatively low inferred from ISSRs (He = 0.028, 0.023, and 0.053 in C. decandra, C. tagal, and C. australis, respectively). No haplotype diversity within population was detected from any of the three species. Cluster analysis based on ISSRs identified three major geographical groups in correspond to the East Indian Ocean (EIO), South China Sea (SCS), and North Australia (NA) in both C. decandra and C. tagal. The cladogram from DNA sequences also detected the same three geographical groups in C. decandra. Analysis of molecular variance (AMOVA) revealed that most of the total variation was accounted for by differentiation between the three major geographical regions of both C. decandra and C. tagal. The significant genetic structure may result from the geological events in these regions during the recent Pleistocene glaciations. This study also provided insights into the phylogenetics of Ceriops. Yelin Huang and Fengxiao Tan contributed equally to this work.