HONEY TYPES AND POLLEN GRAINS OF ASUNCIÓN, LAVALLE, MENDOZA, ARGENTINA (32º33’21’’S/68º14’45’’W), VEGETAL ORIGIN AND POSSIBLE MANAGEMENT OF THE BEEHIVE PRODUCTION

Mónica Cristina Wingenroth

[email protected] Casilla de Correo 330 5500 Mendoza Argentina

Summary This work identifies the species and pollen percentages determined in the selected honey types and pollen grains of beekeepers at Asunción, Lavalle. It also proposes methods of control for honey selection as well as determining the number of individuals (pollen) and the residue in 10 g of the selected honeys.

Key words Pollen, pollen loads or grains, honey, Apis mellifera L., melliferous flora.

Introduction This investigation is warranted by the worlwide interest to encourage organic products for human nourishment and to distinguish the geographical origin of apicultural products through pollen identification, together with the beekeepers’ interest (P.Calderón and family, in this case) to improve the quality of their products, and the environmental urgency to identify economical autochthonous species useful to men. Botanical origin of honey types and pollen grains was achieved as well as the percentage of each pollen species in the beekeepers’ products. The number of individuals (pollen) and residue was also determined in 10 g of honey. Similar studies were approached in nearby phytogeographical regions by Telleria (1992, 1996), Costa and others (1995) and Valle and others (1995). To improve the quality of the products in the area, a control was made of the temporal sequence of pollen appearance in unripe honey types and pollen grains.

Geographic position The present research was done at the Puesto Santo Domingo (32º33’21’’S/68º14’45’’O), located outside of Asunción, Lavalle, approximately between 590-700 m asl (Fig. 1).

Climatology Data from Estrella H.A., Heras V.A., and Guzetta V.A. (1979) belong to Encón (32º15’S/67º50’W, 530 m asl) the nearest meteorological station. The analyzed interval covers the years 1971-1978, showing a mean annual temperature of 18ºC, mean maximum temperature of 25.9ºC, mean minimum temperature of 11.2ºC, absolute maximum temperature of 42ºC, minimum absolute temperature of –9ºC, relative mean humidity of 53%, wind mean velocity of 9.6Km/h, and annual mean precipitation of 103.7 mm.

Proceedings of the 37th International Apicultural Congress, 28 October – 1 November 2001, Durban, South Africa APIMONDIA 2001 To be referenced as: Proc. 37th Int. Apic. Congr., 28 Oct – 1 Nov 2001, Durban, South Africa ISBN: 0-620-27768-8 Produced by: Document Transformation Technologies Organised by: Conference Planners

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ASUNCIÓN El Alpero

ré d n A . G A Lavalle - Mendoza - Argentina

Figure 1. Geographical location of Asunción. (32º33´21´´S /

Vegetation The studied area is included in the “Vegetación de las Travesías”, in the “Mapa de Vegetación de la Provincia de Mendoza” (Roig F., Carretero E. and Mendez E., 1998), specifically in the “Areas de Inundación” with Baccharis salicifolia, Mikania mendocina, Prosopis aff. alpataco, Prosopis flexuosa, Tamarix gallica, among other species.

Material and Methods Plant species of the area were collected and afterwards identified by E.Mendez (IADIZA, CRICYT) and then incorporated in M.Wingenroth Catalogue (IANIGLA, CRICYT). Simultaneously flowering of the species was controlled from October 1999 to April 2000. As in Wingenroth (2000, 2001) flowering was defined as: Initial flowering: stage where the shoots are predominant over the flowers or fruits. Active flowering: stage where the ripe flowers are predominant over the fruits or withered flowers and shoots. Pasive flowering: stage at which the withered flowers are predominant over the shoots or ripe flowers. Pollen was collected from many plants of each species in the area, and treated at the lab (Marcela Marino), using the technique described by Faegri and Iversen (1975), finally mounting the product in silicone oil 2000cst. (Heusser C., personal com.). The slides are included in the Palynological Reference Collection (CRICYT). Honey samples were treated at the lab (M. Marino). To obtain a uniform pollen sample, honey was heated in a warm water bath and stirred, thereafter two samples of 10 g, were taken from each honey. The samples were washed, with distilled and filtered water, centrifuged and decanted. The difference between the empty and filled tube registers the residues weight, after drying the samples, without burning it. To get rid of intersticial water between the micro-particles, glacial acetic acid was added, mixed, centrifuged and decanted. Acetolysis followed, adding 5 ml of a mixture (9 ml anhydric acetic and 1 ml of sulfuric acid) and stirring thoroughly. The mixture was left boiling 2’ in a water bath, always gently stirring, and thereafter centrifuging and decanting. Then 5 ml of glacial acetic acid were added, also centrifuging and decanting. Next the samples were washed with a filtered mixture (7 ml water and 1 ml concentrate detergent), always stirring gently, centrifuging and decanting. Samples were twice washed with tertiary butyl alcohol to get rid of water, in both cases stirring gently, centrifuging and decanting. Then a drop of a mixture (5 ml of tertiary butyl alcohol and 1 ml of silicone) was centered on the slides. All sample was mounted on as many slides as necessary (very carefully to avoid contamination) to make count possible and to obtain a clear sight for identification. Finally the tertiary butyl alcohol was left to evaporate, and the sample was covered and sealed. The temporal sequence of pollen appearance in the unripe honey and in the pollen grains was controlled to improve the quality of honey types in the area, as well as to know the behaviour of Apis mellifera L. This was possible taking unripe honey samples and collecting pollen grains (during 9 consecutive hours) approximately each ten days. Unripe honey samples treated at the lab (D. Calderón and D. Carrasco) were transferred to small precipitation glasses, that were placed in a water bath and left 30’ after boiling started. After cooling, wax was separated and thereafter hermetically stored. The residue, previously mixed, was introduced into clean tubes, distilled and filtered water added, mixed, centrifuged and decanted. Tertiary butyl alcohol was added, mixed, centrifuged and decanted. Procedures follows now, as above described. Acetolysis may be obviate in the treatment of these samples. As mentioned before, approximately each ten days and during nine consecutive hours pollen loads were gathered and their vegetative origin determined. Previously they were selected (M.Marino) by color and size, using for this a Wild-Heerbrugg zoom stereomicroscope. Each pollen grain was imbibed in silicone oil 2000cst, and dispersed with clean needles, finally covered and sealed, and thereafter identified. In the same way 400 pollen grains were mounted, after being randomly selected, from the yearly pollen grain production (1999-2000). Pollen identification was done using the Palynological Reference Collection (CRICYT) in two samples of each analyzed honey (10 g) obtaining the percentages of each species, over a number of at least 2000 individuals, that were also counted in the two samples of each honey. Pollen was also identified in the unripe honey, identifying generally 200 individuals. A Leitz-Dialux microscope was used for the identification of the pollen, generally with 12.5X oculars and 100X objective. Photos were done with a Leitz-Ortholux microscope, with 10X ocular and 16X (general view of honey types and pollen grains) and 100X (predominant pollen species in the honey types and pollen grains) objectives.

Results The vegetation of Puesto Santo Domingo (Asunción) was identified and flowering season followed (T. 1, 2). Pollen was identified and species percentages determined in the honey types of the area (T.3). Pollen species were identified in the pollen loads, and the percentage of this last determined in the product (T.7) Species flowering, together with the temporal appearance of pollen in unripe honey, as in the pollen grains within the area were quoted (T. 2, 4, 8). Pollen number (individuals) in 10 g of each analyzed honey was determined (T.5). Residue weight was registered in each 10 g of the analyzed honey types (T. 6).

Figure 2. General view of the bifloral honey of Capparis atamisquea and Prosopis ( flexuosa or aff. alpataco).

Figure 3. General view of the unifloral honey of Capparis atamisquea

Figure 4. General view of the unifloral honey of

Tamarix gallica

Figure 5. Pollen loads of Capparis atamisquea

Figure 6. Pollen loads of Lycium tenuispinosum

Figure 7. Pollen loads of Prosopis aff. alpataco

Analysis and discussion Comparing the species identified at the area, with the pollen species registered in the honey types, the pollen grains and the samples of the temporal sequence of pollen loads and unripe honey collected, it was observed that Apis mellifera L., visits only some of the autochthonous or naturalized species (T. 1, 3, 4, 7, 8). Taking into account personal studies and data registered by Villagra and Roig (1999) in the vegetation of the area, high percentages of pollen species in the bee products belong to the dominant growing species in the place (T. 1, 3, 7). Following the International Honey Clasification of Louveaux, Maurizio and Vorwohl (1978), honey type I (Fig. 2) frequently registers the presence of Prosopis flexuosa or alpataco, and Capparis atamisquea, rarely the pollen of Bulnesia retama, Prosopis alpataco or flexuosa, Prosopis strombulifera and Tamarix gallica, and sporadically the presence of many other pollen species. Honey type II (Fig. 3) registers very frequently the presence of the pollen from Capparis atamisquea, frequently the presence of the pollen of Tamarix gallica, rarely the presence of Baccharis salicifolia and Prosopis (alpataco or flexuosa) and sporadically the presence of many other pollen species. Honey type III (Fig. 4) records very frequently the pollen of Tamarix gallica, rarely the pollen of Baccharis salicifolia, Capparis atamisquea, Prosopis flexuosa, and sporadically the pollen of many other species (T. 2). Predominant pollen species are recorded only in honeys type II and III, that can be consequently considered of unifloral origin, regarding the authors statements. Prosopis (alpataco or flexuosa) and Capparis atamisquea, are both considered secondary pollen in honey type I, that can consequently be defined as of bifloral origin (T. 3). In the temporal sequence of the unripe honey high percentages of pollen can be observed from Bulnesia retama between the end of November and the beginning of December, Calycera spinulosa at the end of October, Capparis atamisquea starting the 10th of December and ending the 20th of January, Larrea divaricata from the midst to the end of November, Lycium tenuispinosum at the beginning of November, and between the end of February and midst of March, Prosopis aff. alpataco and flexuosa up to the 20th December, Prosopis strombulifera at the end of November, Tamarix gallica between the midst of December to the end of March, generally coincident with the active, sometimes pasive flowering of their species. Data are consequently showing that higher percentages of pollen of Prosopis (flexuosa and alpataco) can be achieved manufacturing only the honey deposited until December 20th in the honeycomb. An unifloral honey of Capparis atamisquea is feasible using only the honey deposited in the honeycomb between the 20th December and January. From the 20th of January on, the unripe honey has very high percentages of Tamarix gallica, with something of Baccharis salicifolia and Lycium sp., giving raise to an unifloral honey of Tamarix gallica. As flowering interval modifies with climatic and environmental conditions, it is wise if the beekeeper controls yearly in the pollen grains or in the unripe honey how and when species pollen percentages of Prosopis (alpataco and flexuosa), Capparis atamisquea and Tamarix gallica, between others, are changing (T. 2, 4). The number of individuals identified in two samples, each of 10 g, in the honey type I, II, III, is between 20.000-100.000, being in this way included in Group II, defined by Maurizio, in Louveaux, Maurizio and Vorwohl (1978), (T.5). But it should be considered the great difference between the number of individuals (pollen) between honey type I-II and III, proposing a subdivision of the Group II of Maurizio as follows, Group II a: 20.000-45.000; Group II: 45.000-100.000. This is also supported by the different taste and color of honey type I-II, belonging to autochthonous species, and honey type III, that belongs to a naturalized species. The residues are similar in the three analyzed honey types (T. 6). From the 400 pollen grains randomly selected, 76% of the population was of unifloral origin (304 pollen loads). In decreasing order the highest percentages are of Capparis atamisquea (Fig. 5), Lycium tenuispinosum (including chilense) (Fig. 6), Prosopis aff.alpataco (Fig. 7), Larrea divaricata, Baccharis salicifolia. Prosopis flexuosa, Prosopis strombulifera which pollen belongs to the species densely represented in the area. Lower percentages, less than 1%, are observed for the pollen loads of Grahamia bracteata, Prosopidastrum globosum, Senecio subulatus, Tamarix gallica, Tessaria absinthioides and Wedeliella incarnata, part probably as response to a low species density within the area, excepting the case of the naturalized Tamarix gallica (Ruiz Leal, 1972), regionally well represented. In the 400 pollen loads identified 22.25% belong to pollen grains of bifloral origin, with Capparis atamisquea-Tamarix gallica highly represented, in spite that Tamarix forms very few unifloral pollen grains. Finally 1.75% of pollen loads are of multifloral origin (T. 7). In the temporal analyzed sequence of pollen loads a very low number of them was collected until 18.10.1999, making the percentages not available. Baccharis salicifolia, Capparis atamisquea, Larrea divaricata, Lycium tenuispinosum (includes chilense), Prosopis aff. Alpataco, Prosopis strombulifera, Tamarix gallica were registered with the highest percentages, while Calycera spinulosa, Prosopidastrum globosum, Prosopis flexuosa, Senecio subulatus, Tessaria absinthioides and Wedeliella incarnata, exhibit very low percentages or not available percentages, being in both cases their presence coincident with the active flowering of the species. The presence of Allenrolfea vaginata, Hoffmanseggia glauca, Lippia nodiflora, Mikania mendocina, Solanum elaeagnifolium and Suaeda divaricata, not randomly selected from the product, agrees with the active flowering of the species and in occasions with the pasive flowering, as in the case of Lippia nodiflora. Bifloral pollen loads are also observed, both species being sometimes involved in passive flowering (Capparis atamisquea-Tamarix gallica), or in active and pasive flowering (Capparis atamisquea- Baccharis salicifolia, Capparis atamisquea-Tamarix gallica, Prosopis aff. alpataco-Prosopis strombulifera), or in active and initial flowering, like in the case of Prosopis aff. alpataco-Prosopis strombulifera. Multifloral pollen grains are also formed, principally during the pasive flowering of the main species visited by Apis mellifera, or at the end of the flowering season, like in the case of the pollen load of Baccharis salicifolia, Hoffmannseggia glauca, Lycium tenuispinosum (includes chilense), Mikania mendocina (T.2, 8). The observed pollen grain production is closely related with the vegetation (density) as per Diaz-Losada and others (1998) and of course with the active flowering of the species. In Asunción, the heaviest pollen grains formed by Apis mellifera belonged to species with reticulate, scabrate, fosulate, striate and also spiny exines (Wingenroth, in elaboration) but always with polar axis and/or equatorial diameter smaller than 30 µ. Pollen with exines characterized by big spines like those of Tessaria absinthioides and polar axis and equatorial diameter bigger than 30 µ form lighter loads, than those of Baccharis salicifolia and Mikania mendocina, both with spiny exines and polar axis and equatorial diameter smaller than 30 µ. This may partially happen because of the work Apis mellifera has to accomplish, filling the holes inbetween the big spines to form the loads, as observed by Vaissière and Vinson (1994) with Gossypium hirsutum (the cotton). Pollen loads formed by pollen species with polar axis and equatorial diameter bigger than 30 µ are generally lighter, as seen in the loads of Grahamia bracteata, Prosopidastrum globosum, Prosopis strombulifera and Wedeliella incarnata, than those loads formed by pollen species with smaller axis and diameter, as the case of Capparis atamisquea, Baccharis salicifolia, Prosopis aff. alpataco subject to be considered when calculating bee diet, as stated by Buchmann and O’ Rourke (1991) (T. 9).

Conclusions 1. Two unifloral and one bifloral honey types were identified in Asunción, Lavalle: • Honey I: Capparis atamisquea and Prosopis (flexuosa or alpataco) • Honey II: Capparis atamisquea • Honey III: Tamarix gallica 2. All analyzed honey types are included in the Group II of Maurizio, proposing a subdivision of this group. 3. Unifloral, bifloral and multifloral pollen loads were identified in Asunción, Lavalle. 4. The three highest percentages of unifloral pollen loads correspond to: • Capparis atamisquea • Lycium tenuispinosum (including chilense) • Prosopis aff.alpataco 5. A flowering control, and pollen identification in unripe honey and/or pollen grains is proposed, for the elaboration and presentation of the honey types in the area. 6. It is observed that the presence of pollen in honey depends on the density of each species in the area, on the flowering season of the species, on the proximity of the combs to the species, and of course on the preferences of Apis mellifera L. 7. Efforts should be made to educate the villagers on the importance of these plants in honey production and induce them to install beehive colonies, which would provide them with an additional source of income. Governments should take notice of this when developing cattle raising or forestry programmes, in areas rich in organic pollen and honey production.

Acknowledgements Specially to both beekeepers Pedro and Daniel Calderón, for their help and advice, and to Francisco Jofré, the owner of the Puesto Santo Domingo (Asunción), for his always kind disposition. To Marcela Marino, Daniel Calderón and Diego Carrasco, to Eduardo Mendez, to Mario Lázaro, to Focuser, to Luis Müller, to Maria Elena Soler and Remedios Marín, respectively, my gratitude for processing samples, determining botanical species, positioning the area, revealing photos, scanning photos, revising English text and preparing figures and tables, respectively. I feel specially indebted to the Mendoza Government (F.I.D.E.S., Alfredo Pereyra) and CONICET for the money received to carry out the proyect. Finally, I wish to thank all members of CRICYT, IANIGLA, IADIZA and ESPACIO APICOLA for their help.

References Buchmann S., O’Rourke M., 1991. Importance of pollen grain volumes for calculating bee diets. Grana 30:591-595 Costa M.C., Decolatti N., Godoy F., 1995. Análisis polínico en mieles del Norte de la Provincia de San Luis (Argentina). Kurtziana 24:133-144 Diaz-Losada E., Ricciardelli-DéAlbore G., Saa-Otero M.P., 1998. The possible use of honeybee pollen loads in characterising vegetation. Grana 37:155-163 Estrella H.A., Heras V.A. Guzetta V.A., 1979. Registro de elementos climáticos en áreas críticas de la Provincia de Mendoza. Cuaderno Técnico 1-79. IADIZA Mendoza Faegri K. and Iversen J., 1975. Texbook of Pollen Analysis. Hafner Press Louveaux J., Maurizio A., Vorwiohl G., 1978. Methods of Melissopalynology by International Commission for Bee Botany of IUBS. Bee World 59:139-157 Roig F., Carretero E., Mendez E., 1998. Mapa de Vegetación de la Provincia de Mendoza. Programa Fitocartográfico Mendocino. Instituto de Investigaciones de las Zonas Aridas (IADIZA) CRICYT Mendoza, Argentina Ruiz Leal A., 1972. Flora Popular Mendocina. Deserta 3. IADIZA Schulz E., Lueke M., 1994. A two year pollen calendar for tradionally produced honey types from Gaya, southern Niger. Grana 33(4-5):254-260 Telleria M.C., 1992. Caracterización botánica y geográfica de las mieles de la Provincia Fitogeográfica Pampeana (Repúblioca Argentina) I: Distrito Oriental. Darwiniana 31(1-4):345-350 Telleria M.C., 1996. Caracterización botánica y geográfica de las mieles de la Provincia Fitogeográfica Pampeana (República Argentina) III: Noreste de la Provincia de La Pampa. Darwiniana 34(1-4):245-249 Valle A.F., Andrada A., Aramayo E., Lamberto S., 1995. Análisis polínico de las mieles del sudoeste de la Provincia de Buenos Aires, Argentina. Invest.Agr.: Prod.Prot.Veg. 10(3) Vaissière B., Vinson B., 1994. Pollen morphology and its effect on pollen collection by honey bees, Apis mellifera L. (Hymenoptera: Apidae), with special reference to upland cotton, Gossypium hirsutum L. (Malvaceae). Grana 33:128-138 Villagra P., Roig F., 1999. Vegetación de las márgenes del Río Mendoza en su zona de divagación (Mendoza, Argentina). Kurtziana 27(2): 309-317 Wingenroth M., 2000. Granos de polen de Asunción (32º33’21’’S/68º14’45’’O), Lavalle, Mendoza, origen vegetal y otras características. Espacio Apícola X(44): 16-29 Wingenroth M., 2001. Granos de polen de Asunción (32º33’21’’S/68º14’45’’O), Lavalle, Mendoza, origen vegetal y otras características. Anexo I. Espacio Apícola XI(41):22-25

TABLE 1 Autochthonous or naturalized species in Asunción (32º33'21''S/68º14'45''W), Lavalle

Species Common name Allenrolfea vaginata (Griseb.)Kuntze jume Amaranthus standleyanus Parodi ex Covas Atriplex vulgatissima Speg.ex Gray zampa Baccharis salicifolia (R.et P.)Pers. chilca Baccharis spartioides (Hook. et Arn.)Remy pichanilla Bulnesia retama (Hook. et Arn.)Gris. retamo Calycera spinulosa Gill.ex Miers barba del león Capparis atamisque O.K. atamisque Cereus aethiops Haw. Chamaesyce catamarcaensis Croiz Cyclolepis genistoides Don Eupatorium patens D.Don ex Hook. et Arn. Euphorbia ovalifolia (Klotz et Garke)Boisser leche palomo Geoffroea decorticans (Gill.ex Hook. et Arn.)Burk. chañar Gnaphalium gaudichaudianum DC. Gomphrena tomentosa (Gris.)Fries hierba del sapo Grahamia bracteata Gill.ex Hook. et Arn vinagrillo Heliotropium curassavicum L. Heliotropium mendocinum Phil. Hoffmanseggia glauca (Ort.)Pfeifert porotillo Ibicella parodii Abbiatti cacho de cabra Larrea divaricata Cav. jarilla Lecanophora heterophylla (Cav.)Krapov. Leptoglossis linifolia (Benth.et Hook.)Gris Lippia nodiflora (L.)Greene Santa María Lycium chilense Miers ex Bertero llaullín Lycium tenuispinosum Miers llaullín Malvella leprosa (Ort.)Krap. Mikania mendocina Phil. enredadera, vejuco Nama undulatum H.B.K. Nicotiana noctiflora Hook. dama de noche Opuntia sulphurea Gillies in Don enmend Schumann Plectrocarpa tetracantha Gill.exHook. Prosopanche americana (R.Brown)Baillon flor de madera, o de piedra Prosopidastrum globosum (Gill.ex Hook.et Arn.)Burk. caballo del diablo Prosopis aff.alpataco Phil. alpataco, lámaro Prosopis flexuosa DC algarrobo Prosopis strombulifera (Lam.)Benth. retortuño Senecio subulatus D.Don ex Hook.et Arn. Solanum euacanthum Phil. Solanum elaeagnifolium Cav. quillo Sphaeralcea miniata (Cav.)Spach. Suaeda divaricata Moq. vidriera Tamarix gallica L. tamarindo Tessaria absinthioides (Hook.et Arn.)DC pájaro bobo Trichloris crinita (Lag.)Parodi Tricomaria usillo (Gill.)Hook. et Arn. usillo Tweedia brunonis Hook.et Arn. Verbena gracilescens (Cham.)Herter Verbesina encelioides (Cav.)Benth.et Hook. Wedeliella incarnata (L.)CKL. roseta del campo

TABLE 2 Flowering interval of the species visited by Apis mellifera L. in Asunción (32º33'21''S/68º14'45''O), Lavalle

Year 1999 2000 Month 10 11 12 1 2 3 Day 8 18 28 9 19 30 10 20 30 10 20 1 9 19 29 11 22 Species Allenrolfea vaginata I I A A P Amaranthus I A standleyanus Atriplex vulgatissima A Baccharis salicifolia I I I A A A A A P P P Bulnesia retama I I A P P P P Calycera spinulosa A A A A A P P P P I A A A P P Capparis atamisquea I I I I A A A A P P P. P P P P. P Euphorbia ovalifolia I A A A P P P P P P Geoffroea decorticans A A P P P P Gomphrena tomentosa A A A P Heliotropium I A A A A A P P P P P A A P P curassavicum Hoffmanseggia glauca I I A A A A A A A A P Larrea divaricata I I I A P P Lippia nodiflora I I I I A A A A A A P P Lycium chilense I A P P A P A A P Lycium tenuispinosum I I I I A P P A P I I A A A P Mikania mendocina I I A A A A A A A A P P Prosopidastrum I I I A A P P P P P P P A A P globosum Prosopisaff.alpataco I I A A A P P P P P P P P P P P. P Prosopis flexuosa I I I A A P P P. P P P P P Prosopis strombulifera I I I A A A P P P A A P P P P Senecio subulatus I A P Solanun elaeagnifolium I I A A A A P P P P Suaeda divaricata P Tamarix gallica I I A A A A P P P P P Tessaria absinthioides I I I A A A A P P Tricomaria usillo I I A A A A A A A A A P P Wedeliella incarnata I I A A A A A A A A A A A

I: initial flowering; A: active flowering; P: pasive flowering

TABLE 3 Pollen species in the honey types of Asunción (32º33'21''S/68º14'45''W), Lavalle Beekeeper: Pedro Calderón and family Harvest: 1999-2000

Pollen of: Honey I % Honey II % Honey III % Allenrolfea vaginata 0.01 0 0.1 Amaranthus standleyanus 0.24 0.1 0 Atriplex vulgatissima 0.08 0.14 0.06 Baccharis salicifolia 0.54 7.9 12.8 Bulnesia retama 9.46 0.1 0.8 Calycera spinulosa 0.91 0.24 1.03 Capparis atamisquea 33.59 49.03 8.02 Euphorbia ovalifolia 0.11 0 0.19 Geoffroea decorticans 0.19 0.1 0 Gomphrena tomentosa 0 0 0.02 Heliotropium curassavicum 0 0.29 0.04 Hoffmanseggia glauca 0 0 0.02 Larrea divaricata 2.96 1.49 0.29 Lippia nodiflora 0.19 0.43 0.7 Lycium chilense 1.69 1.35 0.64 Lycium tenuispinosum 2.41 0.72 0.6 Lycium sp. 0 0 0.41 Mikania mendocina 1.28 0.34 0.48 Prosopidastrum globosum 0.19 0.1 0.08 Prosopis aff.alpataco 8.24 7.23 1.28 Prosopis flexuosa 27.66 1.63 5.73 Prosopis aff.alpataco & flexuosa 35.905 8.86 7.01 Prosopis strombulifera 3.96 0.38 0.97 Senecio subulatus 0.05 0 0 Suaeda divaricata 0.09 0 0 Tamarix gallica 5.74 28.35 65.24 Tessaria absinthioides 0.14 0.05 0.65 Tricomaria usillo 0.02 0 0 Unknown 0.2 0 0.31 Identified pollen number (individuals) 6258 2074 4834

Two different species Prosopis flexuosa or aff. alpataco can be observed, the most likely is mentioned first

TABLE 4 Appearance of pollen species in the unripe honey sequence of Asunción (32º33'21''S/68º14'45''W), Lavalle Harvest 1999-2000 Beekeeper: Pedro Calderón and family

Year 1999 2000 Month 10 11 12 1 2 3 4 Day 28 9 19 30 10 20 30 10 20 1 9 19 29 11 22 4 Pollen of: Hives % % % % % % % % % % % % % % % % Allenrolfea vaginata A B # C Amaranthus standleyanus A 1.92 B C Atriplex vulgatissima A 1.5 B 0 C 0 # Baccharis salicifolia A 1.92 6.06 # # 44.9 # # B 0.62 # # 0.5 2.99 0.19 C 0.66 16.1 13 5.02 1.12 4 0.17 # Bulnesia retama A 2.5 55.6 11.7 B 8.07 60.6 65.5 # 76.8 C 8.95 0.66 35.2 4.96 0.71 # Calycera spinulosa A 1.5 0.68 1.92 # B 67.6 3.72 # C 0.74 0.66 1.06 1 Capparis atamisquea A 10 1.37 1.25 73.5 3.66 38.5 25.8 9.33 2.21 # B 3.72 1.51 9.09 # 27.8 1.27 # 1.49 C 6.71 0.66 7.95 9.22 2.65 # 48.2 # 1 Euphorbia ovalifolia A 1.52 # B # 2.78 C Geoffroea decorticans A 1 B 2.81 C 5.97 1.14 Gomphrena tomentosa A B 0.62 C Larrea divaricata A 4 13.7 7.5 8.67 B 4.34 22.7 10 0.84 C 2.23 1.32 7.95 4.96 1.42 # Lippia nodiflora A B # C 1.14 1.12 Lycium chilense A 6.06 B 8 C 0.66 0.53 5.62 4 Lycium tenuispinosum A 0.4 49 2.74 0.62 0.51 1.92 13.6 2.07 0.44 # # 45.5 B 1.4 6.83 # 3.78 0.91 # 1.69 # 16.4 62 99 # C 5.22 7.28 4.54 4.25 2.66 70.8 25 89.7

TABLE 4 (continued) Appearance of pollen species in the unripe honey sequence of Asunción (32º33'21''S/68º14'45''W), Lavalle Harvest 1999-2000 Beekeeper: Pedro Calderón and family

Mikania mendocina A # 0.44 1.01 B C 0.31 3.37 1 # Prosopidastrum globosum A B 0.62 # C 1.49 0.66 Prosopis aff.alpataco A 0.8 8 38.4 0.62 2.04 22.6 5.77 7.58 # # # B 1.4 47.2 # 5.3 6.36 # 5.55 2.11 2.99 1 1 C 17.9 37.8 10.2 5.67 0.53 # # Prosopis flexuosa A 98.8 38.4 23.1 2.55 51.2 5.77 1.52 1.55 # # # B 8.45 8.69 # 0.75 7.27 # 2.78 13.9 # 4.48 C 40.3 40.4 10.2 63.8 65.4 # # 0.62 2.25 5 Prosopis strombulifera A 1 0.68 1.25 3.05 3.85 3.03 1.04 6.06 B 7.45 # 3.78 0.91 2.78 2.11 # # 3 C 2.98 9.27 21.6 4.25 11.2 Prosopis sp. A 9.37 B C Senecio subulatus A B 3.78 C Suaeda divaricata A 0.5 0.5 B 12.7 0.62 0.75 C 1.49 Tamarix gallica A 14.5 2.74 0.62 1.02 19.5 38.5 40.9 86 # # 52 # # 41.4 # B 1.4 1.24 0.75 # 58.3 1.27 # # # 99.5 71.6 12 0.57 # C 2.23 0.66 2.84 16 # 49.6 # 83.9 86.1 95 15.7 16 10.2 # Tessaria absinthioides A 0.5 B C Unknown A 6 1.37 B 4.22 1.22 C 2.98

# Pollen number (individuals) lower than 50, makes percentages not available

TABLE 5 Pollen number (individuals) in the honey types of Asunción (32º33'21''S/68º14'45''W) Lavalle (10 g) Beekeeper: Pedro Calderón and family Harvest: 1999-2000

Honey I Honey II Honey III 65665 66040 37467 46238 52879 27768

TABLE 6 Residue in the honey types of Asunción (32º33'21''S/68º14'45''W), Lavalle (10 g) Beekeeper: Pedro Calderón and family Harvest: 1999-2000

Honey I Honey II Honey III 0.0388 0.0100 0.0229 0.0162 0.0141 0.0113

TABLE 7 Pollen species in the pollen loads formed by Apis mellifera L. in Asunción (32º33'21''S/68º14'45''W), Lavalle Harvest 1999-2000. Beekeeper: Pedro Calderón and family

Identified pollen loads Percentages

Unifloral pollen loads Baccharis salicifolia 14 3.50% Capparis atamisquea 111 27.75% Grahamia bracteata 3 0.75% Larrea divaricata 35 8.75% Lycium tenuispinosum (includes chilense) 72 18.00% Prosopidastrum globosum 2 0.50% Prosopis aff. alpataco 49 12.25% Prosopis flexuosa 8 2.00% Prosopis strombulifera 5 1.25% Senecio subulatus 1 0.25% Tamarix gallica 1 0.25% Tessaria absinthioides 2 0.50% Wedeliella incarnata 1 0.25%

Bifloral pollen loads Baccharis salicifolia-Senecio subulatus 1 0.25% Baccharis salicifolia-Capparis atamisquea 6 1.50% Capparis atamisquea-Prosopisaff.alpataco 3 0.75% Capparis atamisquea-Prosopis flexuosa 1 0.25% Capparis atamisquea-Prosopis strombulifera 1 0.25% Capparis atamisquea-Tamarix gallica 65 16.25% Larrea divaricata-Prosopis flexuosa 1 0.25% Lycium tenuispinosum-Prosopis aff. alpataco 1 0.25% Lycium tenuispinosum-Wedeliella incarnata 1 0.25% Prosopis aff.alpataco-Prosopis flexuosa 5 1.25% Prosopis aff.alpataco-Prosopis strombulifera 4 1.00%

Multifloral pollen loads Baccharis salicifolia-Calycera spinulosa- 0.00% Capparis atamisquea 1 0.25% Baccharis salicifolia-Capparis atamisquea- 0.00% Prosopis aff.alpataco 1 0.25% Capparis atamisquea-Lippia nodiflora- 0.00% Mikania mendocina-Prosopisaff.alpataco 1 0.25% Capparis atamisquea-Prosopisaff.alpataco 0.00% Prosopis flexuosa-Prosopis strombulifera 1 0.25% Prosopisaff.alpataco, Prosopis flexuosa- 0.00% Prosopis strombulifera 3 0.75% Total pollen loads analyzed 400 100.00%

TABLE 8 Temporal sequence of the pollen loads formed by Apis mellifera L. in Asunción (32º33'21''S/68º14'45''W), Lavalle Harvest 1999-2000. Beekeeper: Pedro Calderón and family Year 1999 2000 Month 9 9 10 10 10 11 11 11 12 12 12 1 1 2 2 2 2 3 3 Day 15 26 8 18 28 9 19 30 10 20 30 10 20 1 9 19 29 11 22 Percentages % Pollen loads with unifloral origin Allenrolfea vaginata 0.02 Baccharis salicifolia 6.56 42.9 16.5 50.8 32.8 2.63 0.14 24.7 Calycera spinulosa 1.84 0.31 0.77 Capparis atamisquea 72.7 57.3 91 87.4 90 13.5 1.85 29.9 0.2 0.59 0.19 Hoffmanseggia glauca 0.13 0.72 Larrea divaricata 22.6 27.3 19.2 0.24 10.3 1.03 Lippia nodiflora 7.02 0.04 3.69 Lycium tenuispinosum (includes chilense) 35.3 25.8 38.5 0.69 6.35 43.7 4.67 7.35 0.82 95.9 99.4 94 70.6 Mikania mendocina 4.28 0.28 Prosopidastrum globosum 58.8 51.6 Prosopis aff.alpataco 62.2 92.8 31.8 4.87 1.16 12.6 1.19 2.21 Prosopis flexuosa 0.14 Prosopis strombulifera 11.5 13.8 3.33 Prosopis sp. 0.11 Senecio subulatus 0.17 Solanum elaeagnifolium 6.4 0.02 Suaeda divaricata 5.88 Tamarix gallica 17.9 60.7 Wedeliella incarnata 1.94 0.41 1.6

Pollen loads with bifloral origin Capparis atamisquea-Baccharis salicifolia 0.83 Capparis atamisquea-Tamarix gallica 0.95 1.67 13 18.9 62.2 Larrea divaricata-Prosopis aff. alpataco 0.39 Prosopis aff.alpataco-Prosopis strombulifera 30.8 36.7 9.38 26.9 0.2 Prosopis aff.alpataco-Prosopis flexuosa 2.91 0.55 Baccharis salicifolia-Prosopidastrum globosum 0.02

Pollen loads with multifloral origin Larrea divaricata-Prosopis aff.alpataco- Prosopis strombulifera 0.56 Prosopis aff.alpataco-Prosopis flexuosa- Prosopis strombulifera 0.08 Baccharis salicifolia-Lycium tenuispinosum- Mikania mendocina-Wedeliella incarnata 0.15 Baccharis salicifolia-Hoffmanseggia glauca- Lycium tenuispinosum-Mikania mendocina 0.02 Number of pollen loads per species 17 31 11 26 2017 1243 2814 2509 7346 2224 721 1750 484 5076 3588 3864 2169 4111 9321 Total number of pollen loads: 49322

TABLE 9 Unifloral pollen loads weight in Asunción (32º33'21''S/68'14'45''W), Lavalle Beekeeper: Pedro Calderón and family Harvest: 1999-2000

Species Number of Weight; mg weighted grains Allenrolfea vaginata 4 5.50 Baccharis salicidolia 50 6.18 Bulnesia retama 3 7.26 Calycera spiunulosa 1 5.50 Capparis atamisquea 152 7.05 Grahamia bracteata 1 4.40 Larrea divaricata 36 7.85 Lycium tenuispinosum 156 6.02 Prosopidastrum globosum 1 4.20 Prosopis aff. alpataco 88 7.12 Prosopis flexuosa 3 6.96 Prosopis strombulifera 9 4.92 Tamarix gallica 20 8.00 Tessaria absinthioides 5 5.88 Wedeliella incarnata 1 6.33