Phenology determines the different phases of the life cycle of a plant and plant productivity. Pollens play an important role in the productivity of a plant. A plant requires sufficient viable pollens for efficient pollination. Meiosis during microsporogenesis in the anther leads to the formation of pollens representing male gametophyte. The pollen grains after pollination on the stigma produce pollen tubes emerge through pollen apertures and then these pollen tubes grow down through the style tissue to fertilize egg and form embryo. The morphology of pollen grains, particularly the openings permitting pollen tube germination (apertures), is crucial for determining the outcome of pollen competition in flowering plants. Numerous studies have been undertaken on pollen morphology in various trees in relation to its taxonomy [1-3]. Thelarge variations observed in pollen morphology with respect to size, shape, surface structure, and germ spores, are of great importance in taxonomic delimitation of species and their evolutionary sequence and also its use in applied fields such as criminology and geology. In this respect, studies have been undertaken on various species viz., Acalypha [4]. Similar studies have been undertaken on the subtribe Nepetinae, similarly on vegetation communities in arid regions of China (Luo et al.), and morphology and ultrastructure of species of Magnoliaceae which depicts an early trend of specialization. This supports the view that Magnoliaceae are not one of the earliest lines in the phylogeny of flowering plants [5]. Kovacik et al. [6] studied the three-dimensional (3-D) structure of channels and bacula cavities in the wall of hazel pollen grains. Sachin et al. studied pollen morphology and pollination biology of Amorphophallu sspecies from North Western Ghats and Konkan of India. Lu et al. [7] studied the pollen morphology of 86 samples from 84 species of Gaultheria and the closely related genera Chamaedaphne, Craibiodendron, Diplycosia, Eubotrys, Gaylussacia, Leucothoe, Lyonia, Oxydendrum, Pieris, Satyria, and Vaccinium (subfamily Vaccinioideae). Using pollen morphological characteristics, a new non-molecular evidence was put forward to support the phylogenetic and taxonomic position, pollen grains of 20 populations of 16 species of Chinese Curcuma L. and Boesenbergia Kuntz (Zingiberaceae) [8]. Similar studies are undertaken by various authors, Khansari et al. studied the pollen grains of 47 taxa of Campanulaceae, Beretta et al. [9] on European bladderworts: Utriculariaaustralis, U. bremii, U. intermedia, U. minor,U. ochroleuca, U. stygiaand U. vulgaris [10] on pollen morphology of Prunusdomestica cv. PožegacaL.

Pollen viability

With respect to the technique Acetocarmine and fluorescein diacetatewas was used to assess the pollen viability of four Požegacaplum ranging from 67% to in 99%. The pollen nucleus was binucleate. The highest pollen germination (96%) and a tube length (822 μm) of genotype PdP4 was observed in media with 20% PEG [10]. Mazzeo et al. [11] used three different techniques for estimating the pollen grain viability: acetic carmine, fluorescein di-acetate and germination. Pollen viability and pollen tubes growth have profound effects on fruit and seed production. Knowledge of pollen biology is therefore, necessary to increase the productivity of a tree. In this context, a concise review of literature has been made herein. A study has been undertaken on meiosis of mandarin on the meiotic behavior and pollen viability in an open-pollinated population of the ‘Lee’ mandarin [Citrus clementina× (C. paradise × C. tangerina)] in Southern Brazil. In most plants the microsporogenesis was regular, with meiotic indexes over 90% and pollen viability over 83%. Several meiotic abnormalities were observed. It is concluded that most plants have the ability to produce viable gametes and could be used as pollen donors in programmed crosses. The monoembryonic unreduced-gamete producer plants are potential progenitors in crosses aimed at producing triploid seedless fruits in mandarin [12]. The environment has effects on the reproductive development and pollen biology of a plant. Sukhvibul et al. [13] reported the effects of temperature on inflorescence and floral development in four mango (Mangiferaindica,L.) cultivars. Day/night temperatures of 15/5°C severely inhibited the emergence and elongation of inflorescences of all cultivars with inflorescence development only occurring on trees that were maintained at warmer temperatures (20/10°C, 25/15°C and 30/20°C). Generally, warmer temperatures increased the inflorescence size of all cultivars. At 20/10°C, the pollen viability of ‘sensation’ was significantly lower than the other cultivars, but there was no significant difference between the cultivars held at 25/15°C and 30/20°C. Huang et al. [14] studied the effect of low temperature on the sexual reproduction process of ‘Tainong 1’ mango at diurnal maximum temperature of <20°C. The results indicated that low temperatures prevailing during reproductive development significantly affected pistil and male gametophyte development, resulting in poor pollen viability owing to few meiotic chromosomal irregularities as univalents, multivalents, laggards, bridges and micronuclei, and higher incidences and significantly greater proportions of nucleolus fragmentation. Pollen tube growth was delayed under low temperature stress. This clearly indicated the need for taking proper case to protect fruiting plants from environmental hazards. Wang et al. [15,16] studied the pollen flow and mating patterns of a subtropical canopy tree (Eurycorym buscavaleriei) in a fragmented agricultural landscape amidst China’s subtropical forests. They determined pollen dispersal and mating patterns were estimated in two physically isolated stands of E. cavaleriei within fragmented forests using six highly polymorphic microsatellite loci. Although substantial amount of pollens travelled less than 100 m, paternity analysis revealed that a large extent of long-distance pollination occurred in the two stands, respectively. It is assumed that high genetic diversity was found in both adults and offspring and habitat fragmentation did not have a negative impact on E. cavaleriei.

A study was undertaken on pollen viability of obeche trees (Triplochitonscleroxylon K. Schum.) obtained from different flowers located in Ibadan (7°17′N and 3°30′E). Pollen viability varied significantly between the flowers from the three sources. Significant differences were also observed in pollen tube growth between the flowers from each of the trees [17].

The viability of pollen was studied in a widely distributed anemophilous species, Scots pine (Pinussylvestris L.), along a south– north gradient from 60 up to 69°N in Finland. The results showed marked variations in pollen viability both among and within latitudes and also among different temperature treatments. Pollen germination at the smaller temperatures was low across all the study areas, but very good at the highest temperature, which was 20°C. The variation in pollen viability among stands might cause differential success of pollen of different origin suggesting the possibility of gene-flow from south to north was assessed (Parantainen and Pulkkinen).

Acacia mearnsii (black wattle) is a commercially important forestry species in South Africa. In-vitro agar media germination tests in ACIAR and Brewbaker and Kwack media and then used together with vital stain tests (Sigma® DAB peroxidase and p-phenylendiamine) on pollen germination and viability of A. mearnsiipollen revealed that vital stain tests had significantly (p<0.05) higher pollen viability than the agar germination tests [18,19]. A study was done on the pollen viability, in vitro pollen germination and in-vivo pollen tube growth in the biofuel seed crop Jatrophacurcas using 2,3,5-triphenyltetrazolium chloride (TTC). Pollen germination was significantly higher in an agar-based medium composed of sucrose, boric acid and calcium nitrate compared with the control and indole-3-acetic acid (IAA) treatment. Pollen tubes from both self- and cross-pollinated flowers entered the ovary within 8 hours after pollination [20]. In an investigation on pollen viability of three pollinizer mango cultivars, under four storage conditions (room temperature, −4°C, −20°C and −196°C), three methods of pollen viability testing, viz., in-vitro germination, fluorescein di-acetate (FDA) and acetocarmine staining were used. Storage methods and interaction between storage methods and days of storage had highly significant effect on pollen viability (p ≤ 0.0001). Irrespective of mango genotypes, cryo-stored (−196°C) pollens showed significantly higher viability as compared to all the other storage conditions. It was assessed that pollen viability assays confirmed that in-vitro germination test was more reliable. The authors suggested storage of pollen at −20°C for pollination among cultivars having non-synchronized flowering in a season [21]. Olive pollination is anemophilous and an adult olive tree can produce large amounts of pollen grains spread in the air during the flowering period. The viability of pollen grains can be checked by using different methods such as cytoplasmic stains, enzymatic reactions or germination. Among the three techniques, acetic carmine gave the highest values but staining also heat-killed pollen grains. The number of pollen grains significantly varied among the cultivars and between the two years. The variation in pollen in pollen viability was observed in between two years [11].

This study was conducted at Linares municipality of the state of Nuevo Leon, northeastern Mexico. The climate is subtropical and semiarid with a warm temperature. Annual mean temperature is 22°Cand rainfall 500 mm. The main type of vegetation is known as Tamaulipan Thornscrub or subtropical Thorn scrub woodlands (INEGI, 1986). Four native species (trees/shrubs) were selected of high economic importance in the semiarid regions of northeast Mexico, namely, Cordiaboissieri,author family “Anacauita”;Parkinsoniatexana “Palo verde”, Cenizo;Parkinsonia aculeate “Retama”r; Leucophyllumfrutescens “Cenizo”.

The scope of the study limits within a brief and selective review of research advances pertaining to general pollen morphology and pollen viability in relation to plant productively. We studied phenology and a general morphology of pollen of these four species. We studied general pollen morphology by squashing the anthers and staining the pollens with 1% safranin in water and observed under 40x. The study on pollen viability started from 20 May up to the end of June, 2014. We collected flowers from 11 A.M. to 12 A.M. This study was undertaken to determine the effect of the prevailing environmental temperatures (23°C, 28°C, 30°C and 32°C), on pollen viability percentage of four species of woody trees, at the time of collection. We adopted two techniques 1) staining with 1% safranin and 3% iodine in KI, the latter is found to be better than the first one. The fresh anthers are separated, squashed and stained with 3% iodine in potassium iodide for 10 minutes and then counted the number of pollen grains stained as viable at 40x under light microscope, each with 10 replications. We calculated average percentage of viable pollens and standard errors and then presented graphically.

All the four species started flowering from March till June, 2014. In the month of June end some of them started fruit maturity viz., Cordiaboissieri, author family “Anacauita”, Parkinsoniatexana “Palo verde”, Parkinsoniaaculeata “Retama”, Leucophyllumfrutescens “Cenizo” (Figure 1).

Figure 1: Flower structure of four species under study.

Pollen morphology

We could not take detailed pollen morphology owing to the nonavailability of the facilities. The general morphology of pollens are depicted and described in Figures 2 and 3. Among the four species studied Cordata boissieri possessed longest (0.42 cm), followed by Leucophyllum frutens (0.33 cm), Parkinsonia aculeate (0.27 cm) and Parkinsonia texana (0.25 cm). In Cordata boissierithe pollen is more or less ovoidal with surface regular, smooth with little ondulations, two visible germ pores. Both sexine and intine are thickwalled. The pollen grain of Leucophyllum frutensis oval with regular, smooth surface, sexine thick walled and intine thin walled, bi-partite, germpore one visible. In Parkinsonia texana the pollen is more or less pentangular in shape, suface shows little ondulations. Sexine is mediumly thickwalled, intine thin walled. In Parkinsonia aculeate, the pollen is more or less round with irregular surface. Sexine is composed of ovoidal cells with surface ondulations, projections. Two to three germ pores are visible. Kovacick, et al. [9] undertook electron tomography of structures in the all of hazel pollen grains

Figure 2: a) Photograph of pollen of Cordia boissieriat 10x , Length of pole 0.042 cm. b) Photograph of pollen of Leucophyllum frutescensat 10x,Length of polen 0.033 cm.

Figure 3: (a) Photograph of pollen of Parkinsonia texana at 10x, length of pollen 0.025 cm. (b) Photograph of pollen of Parkinsoni aaculeata at 10x, length of pollen 0.027 cm.

In the present study the four species showed large variations in pollens in size, shape, surface ornamentations which coincide with the findings of other authors on various plant sspecies [1-3]. Most of these authors use the morphological characteristics in taxonomic delimitations and the evolutionary relationships of the species and few its utility in criminology.

Pollen viability

Table 1 depicts the effects of temperature on pollen viability of 4 woody species. It is observed from the Table 1 the species studied show large variations in pollen viability in response to the prevailing temperatures Depending on the environments we have taken data on percentage of pollen viability of four species viz. C. boissieri “Anacahuita”, P. aculeata “Retama”, P. texana “Palo verde” and L. tennense”Cenizo” at four temperatures. It was observed that at 23°C the pollen viability percentage in all the four species were 80%. With an increase in temperature the responses of 4 species with respect to pollen viability percentage showed variations among them. In the case of Cordia boissieri the pollen viability reduced drastically from 81% to 54% thereby showing susceptibility to an increase in environmental temperature, while in the case of other 3 species, P. aculeata “Retama”, P. texana” Palo verde” and L.tennense’”Cenizo” the pollen viability did not reduce drastically. From 23 to 30 C, the viability percentage in these 3 species is more orless stable/or fluctuating, but at 32 nc, their viability percentage is reduced to some extent except P.texana” Palo verde” showed drastic decrease in pollen viability at 32°C , from 79 to 64%. It is expected that this variability in responses have direct impact in fertility and fruitification (not attempted in this present study). Similar to the present study, the variability in pollen viability in response to temperatures has been documented in various trees. The pollen viability obeche trees (Triplochitons cleroxylon K. Schum) obtained from different flowers located in Ibadan (7°17′N and 3°30′E) showed significant variations between the flowers from the three sources and also significant differences were also observed in pollen tube growth between the flowers from each of the trees [17].

Table 1: Effect of environmental temperatures on pollen viability of four woody species.

Similarly in a study on the pollen viability on anemophilous species, Scots pine (Pinussylvestris L.), along a south–north gradient from 60 up to 69°N in Finlandshowed marked variations in pollen viability both among and within latitudes and also among different temperature treatments. Pollen germination at the smaller temperatures was low across all the study areas, but very good at the highest temperature, which was 20°C. The variation in pollen viability among stands might cause differential success of pollen of different origin thereby, suggesting the possibility of gene-flow from south to north was assessed [22]. It is reported that at low temp. 20/10°C, the pollen viability in few mango cultivars are reduced drastically, while at 25/15°C no significant differences were observed among mango cultivars [13]. In another study low temperature affected pistil and male gametophyte development resulting to poor pollen viability in mango [14]. Low temperature affect pollination in mango (Dag et al.,), pollen germination in Pistaciavera) in vitro pollen germination and pollen tube growth of Pistacia spp. and Almond (Prunusdulcis Mill.) [23]. Akoroda [24] reported longterm storage of yam pollen. Alcaraz et al. [25] studied in vitro pollen germination in avocado (Perseaamericana Mill.) and optimization of the method and effect of temperature.

The results of the present study show large variation in flowering phenology and pollen viability in response to the environments. The present findings support the findings of various authors demonstrating the effects of environments and temperature on pollen viability. Unlike that in mango low temperature less than 20°C reduce pollen viability, the species showed decline in pollen viability with an increase in temperature in Cordia boissieri, though in others showed fluctuation of stable. Pollen vabilty plays an important role in fertility and embryo development. There is a necessity of concerted research input on pollen biology of trees and shrubs in semiarid Northeast Mexico.