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A melissopalynological study of 54 Louisiana (U.S.A.) honeys
Review o f Palaeobotany and Palynology Elsevier Publishing Company, Amsterdam-Printed in The Netherlands
A MELISSOPALYNOLOGICAL STUDY OF 54 LOUISIANA (U.S.A.) HONEYS
MEREDITH HOAG LIEUX 1
Department o f Botany, Louisiana State University and A. and M. College, Baton Rouge, La. (U.S.A.) (Received August 6, 1970) (Resubmitted September 1, 1971)
ABSTRACT Lieux, M. H., 1972. A melissopalynological study of 54 Louisiana (U.S.A.) honeys. Rev. Palaeobot. Palynol., 13: 95-124.
Pollen analyses were made of 54 commercial Louisiana (U.S.A.) honeys collected during 1967-1968. Fifty-eight different pollen types were identified. A direct correlation was assumed between a plant's pollen quantity and its nectar contribution. On this basis, the major Louisiana honey plants that were determined are: Fabaceae, predominantly Trifolium repens (99%), Rubus, Berchemia seandens, and Salix. Plants established as important for their nectar contributions in isolated samples axe: Cephalanthus occidentalis, Sapium sebiferum, miscellaneous Compositae (Asteraceae), Glycine max, and Parthenocissus quinquefolia. Native plants such as Rubus, Berchemia scandens, and Salix contribute more to Louisiana's honey sources than do cultivated plants or introduced plants. Most samples originated from six to fifteen different plant types and thus do not appear to be of particularly diverse botanical origin. The pollen types that indicate late spring, summer, or fall honeys include: Glycine max, Cassiafasciculata, Polygonum, Lagerstroemia indica, Lythrum, Ampelopsis, Trachelospermum difforme, and others. Thirty-one samples could be considered unifloral honeys. Most Louisiana honeys were a clear color and from nectar of flowers and not honeydew. Honeys collected from different regions of Louisiana are not easily distinguished from one another geographically by pollen types except in a few instances. Three pollen types characteristic of Mississippi and Red River floodplain honeys are: Cephalanthus occidentalis, Mimosa strigillosa, and Forestiera aeuminat~ Castanea and llex form a characteristic pollen combination in honeys produced from the pine-oak-hickory uplands and longleaf pine flatwoods of western Louisiana.
1 Present address: Department of Geology, Louisiana State University and A. and M. College, Baton Rouge, La. 70803 (U.S.A.)
96
MEREDITH HOAG LIEUX
INTRODUCTION Microscopical analysis of honey is based on the fact that the raw materials of honey (nectar and honeydew) have certain constituents which remain identifiable in the ripe honey. For the nectar, these are mainly pollen grains from the blossoms producing the nectar. For the honeydew, these constituents are usually algal cells, fungus spores, and fungus hyphae that come from the surface plants of forest trees. Other materials present in honey include living organisms (especially yeasts), insect body parts, soot, and dust particles. The chief objectives of microscopical examination of honey are to assess the value of different plant species as nectar sources; to determine the geographical origin of honey through its pollen spectrum; to estimate the honeydew contributions by means of fungus spores, fungus hyphae, and algae; to diagnose the poisoning of bees from the poisonous plants they visit; to ascertain the season of production; to check for pollution and adulteration of honey. Results obtained from microscopical analysis of honey concern beekeepers, the honey trade, and State Food Control offices. Beekeepers are especially interested in determining the botanical origin of honey. They want to locate their hives so as to fully utilize existing honey sources. The honey trade seeks knowledge concerning the botanical and geographical origin of honey to help in establishing quality standards. State Food Control offices in many countries require that the quality of the product stated on the label conforms to reality. Furthermore, some countries with high production costs legally protect domestic honey. They enforce a compulsory declaration of origin which can only be determined by microscopical analysis of honey. In several European countries (e.g. France, Germany, Sweden, and Switzerland) honey analysis research is progressing rapidly towards locating and defining the native honey types. But in the United States there is need for comprehensive inquiry on the subject. This research is a melissopalynological investigation of 54 commercial Louisiana (U.S.A.) honeys in which the pollen grains of entomophilous plants predominate. Its purpose is to study the honeys in regard to botanical origin, geographical origin, season of production, unifloral versus mixed honey, and floral honey versus honeydew honey. This work is the first major pollen study of Louisiana honeys.
METHODS
Collection and preparation of reference pollen A reference slide collection of Louisiana honey pollen was prepared. Pollen samples and herbarium specimens were collected from many plants; herbarium specimens were identified; pollen samples were processed and mounted. The pollen samples were prepared by a slight modification of the Erdtman (1952) acetolysis technique
A MELISSOPALYNOLOGICAL STUDY OF 54 LOUISIANA (U.S.A.) HONEYS
97
(chlorination was omitted). Brief identification notes pertaining either to the honey plants or their pollen were entered on cards and assembled in a reference card f'de.
Collection and preparation of honey samples A 1967 list of commercial beekeepers in Louisiana was obtained from the Division of Entomology, Louisiana Department of Agriculture and Immigration. Selected commercial beekeepers throughout the state of Louisiana were contacted and requested to collect uncontaminated (not mixed with samples from other locations) commercial type honey samples in their given areas during the fall of 1967; spring, summer, and fall of 1968. Pollen was extracted from a 20 g honey sample by centrifugation following dilution (20 cm 3 water) and heating (45 °C). The recovered pollen sediment was treated according to the acetolysis schedule. Enough glycerin jelly was added to give the acetolyzed honey pollen sediment a 1 cm 3 volume. A permanent reference slide was made of each sample. Temporary slides for counting purposes were made using the following technique: (1) A vial containing the pollen sediment-glycerin jelly mixture was heated until the glycerin jelly was quite fluid. (2) The sample was stirred with a warm glass stirring rod. (3) An eyedropper was warmed and used to place one drop of sample on a warm clean slide. The same eyedropper, thoroughly cleaned after each use, was used to prepare every temporary counting slide. (4) A 22 mm 2 (no. 1 or 0) cover slip was centered over the drop. Care was taken to assure that the sample covered the 22 mm 2 area evenly and that none leaked from the sides. (Gentle warming of the slide helped in spreading the drop).
Examination of honey samples and data collection The species frequency percentages were based upon a count of at least 250 pollen grains per sample. Up to eleven traverses per slide (at predetermined points 2 mm apart) were made under high power (40×). The first 250 grains were identified and counted. A traverse was completed even though a 250 grain count was reached beforehand. Thus, in most instances more than 250 grains were recorded. If grains were too scarce on the first temporary slide, two or more slides were made and counted until the 250 mark was reached. The 1 cm 3 sediment-glycerin jelly mixtures that had grains too abundant to allow accurate counting were diluted with 1-3 cm 3 of glycerin jelly before making the temporary slides. An approximate number of pollen grains per 20 g honey sample was calculated as follows (this number was halved for comparative purposes and represents a 10 g sample in the results): N = total number of grains per 20 g honey sample; n = number of grains counted; A = total area of 22 mm 2 cover slip (484 mm2); a = area covered by
98
MEREDITH HOAG LIEUX
11 traverses (108.9 mm2), diameter of microscopic field was 0.45 mm; k = area adjustment constant (4.3); ?~ = (1/a)A; d = number of drops per cc (24 drops). The formula (N = ?~nd) was used in calculating total grain count of the original 20 g honey sample reduced to 1 cm 3. Adjustments to the formula were necessary, if fewer or more than eleven traverses were made, and if the 1 cm 3 sample was diluted with more glycerin jelly. The classification of Zander (1935) was used in this study to indicate pollen and associated nectar source: dominant pollen (over 45%), secondary pollen (16-45%), minor pollen (1-15%); trace pollen (less than 1%) was added for convenience. A dominant pollen (over 45%) denotes a unifloral honey. A direct correlation was assumed between a plant's pollen quantity and its nectar contribution to the honey. RESULTS Total grain counts were made on each of the 54 honey samples. With this data, the number of samples appearing within certain arbitrarily set ranges of pollen grain totals was determined (Fig.l). Of the 54 honey samples studied, 28 (51.9%) had less than 20,000 grains per 10 g; 12 (22.2%) had 20,000-100,000 grains; 14 (25.9%) had more than 100,000 grains. The unifloral honey samples of Trifolium repens I , Rubus, Berchemia scandens, and Salix were arranged in a Maurizio-type table (Maurizio, 1958, p. 105). The Trifolium repens and the Salix type samples fell into two discrete ranges, while the Rubus and Berchemia types fell into three continuous ranges (Table I). The first three unifloral type
TABLE I Pollen analyses of 29 unifloral 10 g Louisiana honey samples collected through 1967-1968 Dominant pollen types
Percentage
Number of honey samples within the indicated range of amount of pollen grains 20,000 20,000-100,000 2> 100,000
Total number of honey samples
Trifolium repens I Rubus Berchemia scandens Salix
45-92 51-93 49-83 45-53
6 5 4 1
9 9 7 4
2 1 3
3 2 2 -
1Fabaceae, predominantly Trifolium repens (99%). samples (Table I) showed pollen grain number in the lower and higher limits of the selected range. Salix unifloral type did not contain any 2> 100,000 pollen grain samples. 1Trifolium repens will refer throughout the text to Fabaceae, predominantly (99%) T. repens.
A MELISSOPALYNOLOGICALSTUDY OF 54 LOUISIANA (U.S.A.) HONEYS
99
29
E
E
2
14 12
( 20,000
20,000-100,000
7100, O00
Pollen grains in 10g honey samples Fig.1. Frequency of honey samples in the range ~ 20,000; (20,000-100,000); ~ 100,000 total pollen grains per 10 g sample.
Respectively 66%, 55%, and 25% of Trifolium repens, Rubus, Berchemia scandens, and Salix samples showed < 20,000 pollen grains in 10 g of honey. Fifty-eight pollen types were identified in the 54 honey samples (Fig.2). (Photographs of most of the 58 pollen types are included in Hate I-VIII). The pollen grains were determined to the family, genus, or species (when possible). Percentage frequencies of the pollen types were calculated and shown in a bar graph (Fig.2). Eight pollen types appeared in 50% or more of the honey samples. In descending order, they are Berchemia scandens, Rubus, Trifolium repens, Salix, Rhus radicans, Rosaceae, Quercus, and Gleditsia. Thirty pollen types appeared in 10% or less of the samples. Fig.3, a Louveaux and Vergeron-type graph (Louveaux and Vergeron, 1964, p. 340), shows a straight-line trend and gives a true picture of Louisiana honey origin. The number of different pollen types per sample is shown (Fig. 4). Of the 54 samples studied, 75.9% had six to fifteen pollen types; 22.2% had sixteen to twenty-five pollen types; and 1.9% had one to five pollen types.
MEREDITH HOAG LIEUX
100
Pollen types
0
IO
20
30
60 50 40 Percent frequency
70
80
Fig.2. Percent frequency of 58 pollen types in 54 Louisiana honey samples.
Pollen spectra by percentages for the 54 samples were grouped according to modifications of Oertel’s (1955) beekeeping areas that are outlined in Fig.5. (These areas were determined on the basis of geology, soil, topography, and vegetation differences). The pollen spectra by percentages are given in Table U-VI. Composite pollen spectra for areas 1, 2,3,4, 5, and for the state are shown in Table VII.
90
100
A MELISSOPALYNOLOGICAL STUDY OF 54 LOUISIANA (U.S.A.) HONEYS
58
55 ,r. ~. z. :~ = _ f . - . . . .
50
~
45
g ~. 4o ~ 35 E ~, 30
Number of honey samples (54)
Fig.3. Straight-line trend representing 54 Louisiana honey samples as to the 58 identified pollen types.
Q_
E
x:l
E
3
1
I
3 (I-5)
8 (6- I0)
13 (11= 15)
18 (16-20)
23 +- P.- Type Average (21-25) (- P.- Type Range
Number of pollen types (58)
Fig.4. Histogram showing range of different pollen types within honey samples.
101
MEREDITH HOAG LIEUX
102
PLATEI
I
0
E
G
I( |
!
"50u
Pollen grains representing pollen types identified in the boney samples studied. A. Allium canadense L. (106-104A). B,C. Ampelopsis arborea (L.) Koehne (160-160A). D,E. Berchemia scandens (Hill) K. Koch (111-109A). F,G. Brassica (31-24A). H. Brunnichia cirrhosa Gaertn. (166-169A). J,K. Callicarpa americana L. (25683-238A). Magnification X 900. Code number refers to the herbarium specimen and the reference pollen slide in the Lieux collection.
A MELISSOPALYNOLOGICAL STUDY OF 54 LOUISIANA (U.S.A.) HONEYS
103
P L A T E II
Oi) I
L
•
!
,50u
Pollen grains representing pollen types identified in the honey samples studied. A,B. Cassiafasciculata Michx. (188-192A). C,D. Castaneapumila (L.) Mill. (404064-236A). E,F. Celtislaevigata Willd. (73-87A). G,H. Cephalanthus occidentalis L. (158-157A). J. Cocculuscarolinus (L.) DC. (Brown 1261). K,L. Cornusdrummondi Meyer (Brown 775). M,N. Cornusflorida L. (99-59A). Magnification X 900. Code number refers to the herbarium specimen and the reference pollen slide in the Lieux pollen collection.
104
MEREDITIt HOAG LIEUX
P L A T E III
4
8
I |
,( it ,
,,,
,50U
Pollen grains representing pollen types identified in the honey samples studied. A,B. Forestiera acuminata (Michx.) Poir. (462328-232A). C,D. Fraxinusamericana L. (171488-233A). E,F. Gleditsia aquatica Marsh. (Brown 1/61). G,H. Gleditsia triacanthos L. (137723-227A). J,K. Glycine m a x (L.) Merr. (196-194A). L,M llex cornuta Lindl. (57-55A). Magnification X 900. Code number refers to the herbarium specimen and the reference pollen slide in the Lieux pollen collection.
A MELISSOPALYNOLOGICAL
STUDY
OF 54 LOUISIANA
(U.S.A.)
JIONEYS
105
PLATE IV
I
Pollen A. B,C. D. E. F. G. H. J,K. L,M.
ISOU
I
grains representing pollen types identified in the honey samples Lagerstroemia indica L. (Brown 1585). Ligustrum (154-152A). Liquidambar styraciflua L. (46A). Lythrum alatum Pursh. (190-190A). X 450. Magnolia grandiflora L. (154-153A); magnification Mikanea scandens (L.) Willd. (Brown 1375-18761). Mimosa strigillosa Torrey et Gray (186-165A). Nyssa sylvatica Marsh. var. biflora (Walt.) Sarg. (Brown 746). Parthenocissus ipinquefolia (L.) Planch. (335458-219A).
Magnification X 900, unless otherwise stated. Code number reference pollen slide in the Lieux pollen collection.
studied.
refers to the herbarium
specimen
and the
106
MEREDITH HOAG LIEUX
PLATE V
,,L
C E
IP
U
It
1
i
,50u
Pollen grains representing pollen types identified in the honey samples studied. A. Planera aquatica (Walt.) J.F. Gmel. (99318-231A). B. Polygonum (Brown 1226-18541). C,D ,E.Prunus caroliniana Mill. (27-20A). F,G. Prunuspersica (L.) Batsch (33-23A). H. Prunus serotina Ehrh. (63-37A). J. Pyrus (20-15A). Magnification X 900. Code number refers to the herbarium specimen and the reference pollen slide in the Lieux pollen collection.
107
A MELISSOPALYNOLOGICAL STUDY OF 54 LOUISIANA (U.S.A.) ItONEYS PLATE VI
B
E
i,
F
G
L
K tt
M
V
!
I
,50u
Pollen grains representing pollen types identified in the honey samples studied. A. Quercus rnichauxii Nutt. (Brown 774). B,C,D. Rhus radicans L. (Brown 963-17728). E,F,G. Rubus (40-36A). Rumex crispus L. (Brown 759). H,J. K,L,M. Salix nigra Marsh. (23-16A). N,O,P. Sapium sebiferum (L.) Roxb. (Brown 1584). Magnification X 900. Code number refers to the herbarium specimen and the reference pollen slide in the Lieux pollen collection.
108
MEREDITH HOAG LIEUX
PLATE VII
A
l
E V
,d•
H
wll
,,
,
,
.......
, 50U
Pollen grains representing pollen types identified in the honey samples studied. A,B. Senecio glabellus Poir. (44-44A). C. Trachelospermumdifforme (Walt,) Gray (446838-206A). D,E. Trifoliurn incarnatum L. (65-79A). F,G. Trifolium repens L. (86-73A). H. Ulmusfulva Michx. (Brown 762). J,K. Verbenarigida (L.) Spreng. (Brown 1244-18587). Magnification X 900. Code number refers to the herbarium specimen and the reference pollen slide in the Lieux pollen collection.
A MELISSOPALYNOLOGICAL STUDY OF 54 LOUISIANA (U.S.A.) HONEYS
109
PLATE VIII
r.
A
!1
t,,,
Pollen A,B. C,D. E,F.
D
I
'50u
grains representing pollen types identified in the honey samples studied. Vicia ludoviciana Nutt. (95-63A). Vitis (Brown 729). Zea mays L. (Brown 1290);magnification X 450.
Magnification X 900, unless otherwise stated. Code number refers to the herbarium specimen and the reference pollen slide in the Lieux pollen collection.
I
• 106
60
Fig. 5. Beekeeping areas of Louisiana.
S C A LE
LOUISIANA
55 "56
9= ~.git.4,
we.t
.I
/
I
C ~ n~
k 91
Pine-Oak-Hickory Pine-Oak-Hickory Uplands and Longleaf Pine Flatwoods Prairie Longleaf Pine Flatwoods and Upland Hardwoods Marsh Lands
Area Q Area Q Area Q Area O Area G
i
*Small numbers refer to sampling sites.
Uplands
Bottomland Hardwoods and Cypress
Beekeeping Areas of Louisiana (USA)
Area O
Figure 5.
c
© > o
~D
A MELISSOPALYNOLOGICAL STUDY OF 54 LOUISIANA (U.S.A.) HONEYS
111
TABLE II Percentages of pollen types for honey samples from area 1
Taxa~ples
Acer Allium Ampelopsis Berchemia scandens Brassica Brunnichia cirrhosa Cassia fasciculata Castanea Celtis Cephalanthus occidentalis Cercis canadensis Chenopodium Cocculus carolinus Compositae Comus Cruciferae Diospyros virginiana Forestiera acuminata Fraxinus Gleditsia Glycine max Gramineae llex Leguminosae (Trifolium repens) L igu s tru m Liquidambar styraciflua Lythmm Mimosa strigillosa Nymphaea Nyssa Parthenocissus Pinus Planera aquatica Polygonum Quercus Rhus radicans Rosaceae Rubus Rumex Sagittaria Salix Sapium sebiferum Scrophulariaceae I (?) Trachelospermum difforme l Tentative identification.
10
11
14
17
18
26
31
27
0.4 0.8
12.1
0.9
28.5
1.1
5.9
0.4 2.6
6.7
0.9 0.4 29.0
3.1
1.0
0.4 0.4
0.4
1.7
0.4
0.4 3.1
0.4
0.4
0.4 0.4 0.4 0.8 9.0
66.3
1.6
1.1
6.7
0.4 4.7
0.4 1.5
1.8
2.2 0.4 42.1
1.2 7.8 3.9 7.0
1.1 1.1 3.4 1.1 4.1
9.0
48.9
39.6
18.4 0.9 16.0 0.4
0.7
1.1 2.7 1.1 38.9
41.2
0.4 10.3 3.9 21.3 0.4 O.7 30.9
0.9
50.9
3.0
43.8
7.9
0.9
0.4 1.1
0.4 0.7 44.0 0.4 0.4 10.8
0.7
12.6
0.4 1.5
0.4
17.5 1.1
0.4
1.9 3.7 1.1
4.1
MEREDITH HOAG LIEUX
112 TABLE 1I (continued)
•
Samples
10
11
14
17
18
26
27
0.4
2.7
1.1
31
Taxa
Trifolium incarnatum Ulmus Umbelliferae
Verbena Vicia Vitis
1.1
0.9
1.7
0.4 0.7
0.4 0.4
2.4 2.4 0.4 1.2 0.8
44
46
51
62
63
64
1.7
6.4
1.6
20.3
1.4 10.4
4.2 6.5
0.7
UK-11 UK (Unknown) UI (Unidentifiable) mples
A cer Allium Ampelopsis Berehemia seandens Brassiea Bnmniehia cirrhosa Cassia faseiculata Castanea Celtis Cephalanthus occidentalis Cercis canadensis Chenopodium Cocculus carolinus
0.4 0.8
0.4
33
43
0.4
5.2 0.4
1.5 1.1
0.4 1.9
1.9
1.9
3.8 0.6
0.4
Compositae
0.4
Comus
0.4
1.2 0.4
0.3 1.5
1.1 0.7
0.9
0.4
4.9
1.8 6.8
0.9
0.7
42.1
37.4
Cruciferae
Diospyros virginiana Forestiera acuminata Fraxinus Gleditsia Glycine max Gramineae
3.8
0.5
3.8
1.2
flex Leguminosae (Trifolium repens) Ligustrum Liquidambar styraciflua Lythrum Mimosa strigillosa Nymphaea Nyssa Parthenoeissus Pinus
15.0
82.1
13.8
40.6
66.0 0.4
21.3
0.4 0.8
6.2
1.5 0.3
0.4 0.7
0.6
113
A MELISSOPALYNOLOGICAL STUDY OF 54 LOUISIANA (U.S.A.) HONEYS TABLE II (continued)
•
Samples
33
43
44
46
51
62
63
64
0.9 0.5 75.1
8.0 0.4 32.2
3.0 0.8 9.0
0.3 4.0 0.6 0.9
6.1 1.8 3.6
7.4
5.9 0.2
10.0
5.1
15.2
24.1
53.4
Taxa
Planera aquatica Polygonum Quercus Rhus radicans Rosaceae Rubus Rumex Sagittaria Salix Sapium sebiferum Scrophulariaceae (?) Trachelospermum difforme Trifolium incarnatum Ulmus Umbelliferae Verbena Vicia Vitis UK-11 UK UI ~ Samples Taxa " ~
Acer Alliurn A mpelopsis Berchemia scandens Brassica Brunnichia cirrhosa Cassia fasciculata Castanea Celtis Cephalanthus occidentalis Cercis canadensis Chenopodium Cocculus carolinus Compositae Comus Cruciferae Diospyros virginiana Forestiera acurninata Fraxinus
0.4 8.6 34.0 0.8
15.0
3.5
3.8
1.5
0.3
1.9 0.4
0.4
1.1
0.4
1.6 9.4
2.1
0.9 0.3
1.9 0.4 3.8 1.9 65
0.8 1.9 66
0.8 1.6
1.4 67
68
71
2.1 0.3 1.8 72
2.2 73
1.8 75
0.5
42.6
8.6 O.8
52.7
26.3
6.9
39.2
10.0
0.6
0.2 0.9
O.2 5.8
0.7 0.3
0.3
0.3
0.4
2.5 0.5
0.4
1.0
0.3
0.5 0.5
0.2
O.2 0.3 0.3
114
MEREDITH HOAG LIEUX
TABLE II (continued)
Taxa~amples
65
66
67
68
Gleditsia Glycine max Gramineae llex Leguminosae (Trifolium repens) Ligustrum Liquidambar styraciflua L ythrum Mimosa strigillosa Nymphaea Nyssa Parthenocissus Pinus Planera aquatica Polygonum Quercus Rhus radicans Rosaceae Rubus Rumex Sagi t taria Salix Sapium sebiferum Scrophulariaceae(? ) Trachelospermum difforme Trifolium incarnatum Ulmus Umbelliferae Verbena Vicia Vitis UK-11 UK UI
3.6
5.8
2.0
0.7
15.5
0.3 1.0
0.4 0.4 43.6
~
les
Acer Allium Ampelopsis Berchemia scandens Brassica Brunnichia cirrhosa Cassia fasciculata Castanea Celtis
0.3 16.7
71
72
73
75
0.3
0.7
2.5
0.2 23.5
14.9
54.8
0.2 45.4
0.2 11.6
0.9 1.2
0.2
0.8
0.4
0.9 1.8 3.2 ",.-,~
19.0 0.3
1.9 9.3 1.2 35.4
0.5 3.0 8.5 13.4
0.4 15.2
13.4
0.3
0.6 O.3 0.9 0.3 0.3
0.3 0.9 26.3
16.9
5.4
0.5 7.3 0.9 2.7 0.2
3.7 1.3 7.8
17.9
18.2 0.2
0.7
0.2
0.2 0.2 0.2 0.5
1.0
0.4 0.4
0.8
76
79
85
67.4
3.7
27.7
0.3
3.8 2.1 59.2
0.3
0.8 2.3
0.7
3.3 0.7 16.9 0.4
1.5 86
33.2
0.4 1.4
0.3
87
93
107
11.4
12.8
14.4
7.6 0.4
0.2
0.4 0.5 0.3 0.3 0.2
A MELISSOPALYNOLOGICAL STUDY OF 54 LOUISIANA (U.S.A.) HONEYS
115
TABLE II (continued)
Samples
76
79
85
86
87
93
107
Taxa
Cephalanthus occidentalis Cercis canadensis Chenopodium Coeeulus earolinus Compositae
0.3
1.9
0.3 0.4 0.7
Cornus
0.3 0.3
0.4
0.3
6.1
Cruciferae
Diospyros virginiana Forestiera acuminata Fraxinus Gleditsia Glycine max
1.0 0.7
0.8 0.3 0.5
Gramineae
Ilex Leguminosae (Trifolium repens)
Ligustrum Liquidambar styraeiflu Lythrum Mimosa strigillosa Nymphaea Nyssa Parthenocissus Pinus Planera aquatiea Polygon u m Quercus Rhus radieans
13.3
0.4 0.4 6.8
0.3 18.7 0.3
0.4 0.4
6.0
5.9
0.3
0.3 0.2 11.8
1.7
5.3
0.6 1.0
1.3
0.3 6.7
7.6
1.5
0.3 O.8
Rosaceae
Rubus Rumex Sagittaria Salix Sapium sebiferum
56.4
7.9 2.3 1.2
0.6 0.6 1.8 0.3 3.8 2.1 0.9
O.3 O.3
0.3 4.8 2.7 2.7
0.4 2.4 5.2 35.4
0.6 7.6 2.0 28.3
0.8 3.0 1.1 13.6
0.4 8.1 3.8 41.5
2.6 3.8 17.4 19.8
11.0 0.3
17.1 0.8
1.7 1.2
45.4
24.3 0.4
18.4
Serophulariaceae (?)
Traehelospermum difforme Trifolium inearnatum Ulmus
2.3
Umbelliferae
0.5
Verbena Vieia Vitis
O.3
UK-11 UK U!
O.3
1.3
0.3 0.3 0.3 0.5
0.3
0.4
0.3
0.8 0.4 0.8
0.2
0.3 0.3
0.6
0.9
116
MEREDITH HOAG LIEUX
TABLE 1II Percentages of pollen types for honey samples from area 2
Taxa
Sample 30
A Ilium 1.0 Berchemia scandens 41.1 Brassica 0.4 Castanea 3.1 Cephalanthus occidentalis 3.5 Comus 0.4 Diospyros virginiana 0.7 Leguminosae (Trifolium repens) 13.2 Ligustrum 0.4 Magnolia 1.0 Mimosa strigillosa 0.7
Taxa
Sample 30
Parthenocissus Quercus Rhus radicans Rosaceae Rubus Salix Sapium sebiferum Verbena Vitis UI
13.2 0.4 2.4 2.1 3.5 4.5 1.4 1.4 4.2 1.4
TABLE IV Percentages of pollen types for honey samples from area 3
Taxa~
arnples
Berehemia seandens Brassica Brunniehia cirrhosa Castanea Celtis Compositae Comus Diospyros virginiana Fraxinus Gleditsia Gramineae Ilex Lagerstroemia indica Leguminosae (Trifolium repens) Ligustrum Liriodendron tulipifera Magnolia Myrica Nyssa Parthenocissus Pinus
41
55
56
57
60
61
84
29.4
26.6
37.9 O.4
49.8
81.7
28.0
56.3
4.5 6.0
10.0
9.2
5.5
4.0
1.7
4.4 0.4 0.4
0.4
0.4
1.0 0.4
0.4 0.4 1.1
1.9 3.0 6.0
3.5
0.4 1.0
0.7
4.4
0.7 3.8
3.5
4.5
0.3 0.4 0.4 0.4 3.4 17.0 0.4
0.3
1.5
14.4
0.4 0.4
2.8
1.4
2.6
117
A MELISSOPALYNOLOGICAL STUDY OF 54 LOUISIANA (U.S.A.) HONEYS TABLE IV (continued)
Samples
41
55
0.4 0.7 0.7 2.3 12.0 0.4 3.4
5.9 1.0 5.5
56
57
60
61
84
Taxa Polygonum Quercus Rhus radicans Rosaceae Rubus Sagittaria Salix Sambucus Sapium sebiferum Scrophulariaceae (?) Trifolium incarnatum Vitis Zea mays UK-11 UK UI
45.9
0.4 15.2 25.9 6.4 0.4
0.4 6.3 1.1 4.1
0.8
29.1
56.8
11.1
1.7
0.7 1.1
0.7 5.6 0.4 0.4 2.7 0.4 1.9
1.0
0.4
0.4
0.4
0.4 1.0
0.7
2.0
1.0
1.1
TABLE V Percentages of pollen types for honey samples from area 4
T~a~amples
35
Berchemia scandens 37.5 Cassia fasciculata Castanea Compositae 1.9 Comus Fraxinus Gleditsia 0.4 Glycine max Gramineae llex Leguminosae (Trifolium repens) 12.1 Ligustrum Magnolia Parthenocissus Quercus Rhus radicans 8.3 Rosaceae 11.7
38
59
69
0.7 2.9
8.6
8.7
5.0
0.6
1.2 0.8
0.3
29.4
2.8
42.3 0.8
89
20.8
106
6.3
0.5 0.3 0.8 2.0
0.4
40.5 0.4 46.0
70
0.8
0.3
91.6
20.2
5.0 3.1 0.5 2.7 0.9
0.7 1.2
0.4 0.4 3.2 2.3
0.3 12.1 2.2
1.3 0.5
118
MEREDITH HOAG LIEUX
TABLE V (continued)
Samples
Rubus Salix Sapium sebiferum Trifoliurn incarnatum Ulmus Verbena Vitis Zea mays UK UI
35
38
59
69
70
89
106
15.9 9.5 0.4
0.4 0.4
50.7 8.1
20.1 3.2 13.4
3.6
25.6 9.0 2.5 1.4
6.8 2.2 65.6
0.4 0.4
0.7 0.4
0.1
1.1
0.7 0.4 1.5
3.2
0.2
0.8
0.9 0.5
0.3 1.4
1.4 1.8
100
102
103
49.3
83.4
16.3
0.8
TABLE VI Percentages of pollen types for honey samples from area 5
T a x a ~Samples
Berchemia scandens Callicarpa americana Chenopodium
37
2.7
97
98
21.1
27.8
99
2.8
0.3 2.1 86.6
Compositae
Comus Fraxinus
27.1
0.8
0.7 0,4
Gramineae
llex Leguminosae (THfolium repens) Ligustrum Magnolia Nyssa Parthenocissus Quercus Rhus radicans
0.4 1.5 0.4 0.8
0.4 2.2
0.7 0.7 0.3
1.4
4.8 0.4 0.4
1,1 3.3
2.2
0.4
0.4 0.3 2.5 0.1
0.4
1.2 0.8 3.1
4.2 1.9
60.1
73.5
1.5 0.4 61.5
1.1
0.4
3.3
Rosaceae
Rubus Sagittaria Salix Sapium sebiferum
112
0.4 0.3 92.8
19.7
6.6
11,3 8.5
0.5 76.7
3.8 0.1
0.7
Scrophulariaceae (?)
0.5
Trifolium incarnatum Vicia Vitis
2.7
UK UI
0.4 0.8
0.4 1.3 0.7 0.4
1.1
1.4 0.7 0.3
2.8 2.8
0.7 1.1
0.4
0.1 0.2
A MELISSOPALYNOLOGICAL STUDY OF 54 LOUISIANA (U.S.A.) HONEYS
119
TABLE VII Composite pollen spectra by percentages for honey samples from the five main honey producing areas of Louisiana, plus a composite state spectrum
Taxa
Area 1
Acer Allium Ampelopsis Berchemia scandens Brassica Brunnichia cirrhosa Callicarpa americana Cassia fasciculata Castanea Celtis Cephalanthus occidentalis Cercis canadensis Chenopodium Cocculus carolinus
0.01 0.07 0.2 15.2 0.02 0.02
Compositae
Comus Cruciferae
Diospyros virginiana Forestiera acuminata Fraxinus Gleditsia Glycine max Gramineae
Rosaceae
Area 3
Area 4
Area 5
41.1 0.4
44.2 0.05 0.6
0.4
0.2
0.8
12.2
25.4
0. 03
0.01
3.1
0.2 1.5 0.05 0.03 0.01 0.6 0.1 0.03 0.04 0.4 0.3
0.05 3.5 0.3
0.4
0.2 0.1
0.7
O.1
1.4 2.0 0.3 0.1
3.2 17.9
5.8
13.2 0.4
1.3 0.04
0.05
0.1
0.2
0.4 6.5 0.7 0.07
0.2 2.4 0.4 2.0 2.1
10.8 3.6
0.05
0.1 1.0
34.9 0.1
1.5 0.05
0.1
0.06
0.05 0.05
0.5 0.2
0.05 1.0 0.7
0.05
State
0.1 0.06 0.1 20.6 0.02 0.1 0.01 0.2
1.0
0.3
Ilex Lagerstroemia indica Leguminosae (Trifolium repens) 26.1 Ligustrum 0.02 Liquidambar styraciflua 0.02 Liriodendron tulipifera Lythrum 0.01 Magnolia Mimosa strigitlosa 1.6 Myrica Nymphaea 0.01 Nyssa 0.2 Parthenocissus 1.7 Pinus 0.01 Planera aquatica 0.03 Polygonum 0.04 Quercus 0.2 Rhus radicans 4.5 Rub'us
Area 12
0.05
0.1 0.7 0.03 0.06 0.01 2.1 0.6 0.02 0.06 0.2 0.2 0.9 2.0 0.3 0.5 0.06 20.3 0.3 0.01 0.01 0.01 0.05 1.0 0.01
0.01 13.2
0.4 2.4 2.1 3.5
1Area 2 is represented by only one honey sample.
1.6 2.5 0.05 0.05 0.2 4.0 0.6 16.6
0.3 3.5 2.5 17.6
0.6 0.9 0.08 49.3
0.4 1.6 0.1 0.02 0.03 0.5 3.7 2.3 22.1
MEREDITH HOAG LIEUX
120 TABLE VII (continued)
Taxa
Area 1
Rumex Sagittaria Salix Sambucus Sapium sebiferurn Scrophulariaceae (?) Trachelospermum difforme Trifolium incarnatum Ulmus Umbelliferae Verbena Vicia Vitis Zea mays UK-II UK UI
0.03 0.03 17.9
Area 2
4,5
O.1 0.1 0.01 0.09 0.3 0.01 O.2 0.8 0.3
1.4
4.2
0.2 0.4 1.0
1.4
Area 3
0.05 12.5 0.05 O.2 0.1 0.8
1.4
Area 4
Area 5
4.6
0.01 2.1
11.6
1.1 0.06
0.2 0.2
0.4
O.7 0.4 0.05 0.4 0.1 1.1
0.2 0.1 0.3 1.0
0.2 0.6 0.2 0.9
DISCUSSION This research presents information on Louisiana commercial honeys concerning botanical origin, geographical origin, unifloral versus mixed honey, floral honey versus honeydew honey, and season of production.
Botanical origin Botanical origin reveals that the leading Louisiana honey plants are TriJbliurn repens, Rubus, Berchernia scandens, and Salix. They are the major nectar contributors to the honey. Trifolium repens, Rubus, Berchemla scandens, and Salix also have the highest frequency of appearance in Louisiana honey samples. Miscellaneous Compositae (Asteraceae), Sapium sebiferum, Glycine max, Parthenocissus quinquefolia, and Cephalanthus occidentalis are confirmed as important nectar contributors in isolated samples. Miscellaneous Rosaceae (except Rubus) and Rhus radicans are notable for their frequency of appearance. Four plants worthy of recognition did not appear in Oertel's (1955) most recent list of Louisiana honey plants: Sapium sebiferum, Glycine max, Parthenocissus quinquefolia, and Rhus radicans. Sapium sebiferum is an introduced tree. The results indicate that Sapium sebiferurn is now widespread, especially in south-western Louisiana. Increased soybean production could easily account for Glycine max pollen appearing in the pollen spectrum. The Louisiana State Department of Agriculture (1966) stated that the soybean industry has made much progress in the last ten years;
State 0.02 0.03 12.9 0.01 1.8 0.09 0.01 0.2 0.2 0.01 O.2 0.5 0.4 0.02 0.2 0.3 !.0
A MELISSOPALYNOLOGICALSTUDY OF 54 LOUISIANA(U.S.A.) HONEYS
121
in 1965 there were 622,000 acres (more than any other crop) of soybeans grown in Louisiana. It is probable that Rhus radicans was not reported in 1955 because commercial beekeepers hesitated to admit that poison ivy is significant among the nectar-secreting flora. Other kinds of plant pollen appearing in the samples but not in Oertel's (1939) list of honey and pollen plants of Louisiana are: Allium, Chenopodium, Cocculus carolinus, Gramineae (Poaceae), Liriodendron tulipifera, Magnolia, Mimosa strigillosa, Myrica, Pinus, Rumex, Sagittaria, Sambucus, Scrophulariaceae, Umbelliferae (Apiaceae). All except Mimosa strigillosa generally appeared as trace pollen (less than 1%). It is assumed that the pollen grains from wind-pollinated plants such as Gramineae (Poaceae), Rumex, Pinus, Quercus, and Sagittaria are accidental honey contaminants. The pollen grains may have fallen or been blown into the nectar or honey; or were deliberately collected by the bees, stored in pollen cells, and entered the honey during the extraction process. Louisiana plants reported as honey or pollen plants by Oertel (1939), but whose pollen did not appear in the honey samples are: Ailanthus altissima, Aralia spinosa,
Bumelia, Citrus, Cliftonia, Convolvulus, Corculum leptopus, Cyrilla, Gossypium, Halesia, Ipomoea, Jacquemontia, Jussiaea, Melia azedarach, Oxydendrum arboreum, Perilla frutescens, Phoradendron flavescens, Richardia scabra, Sabal minor, S tellaria media, S tizolobium deeringianum, Tecoma radicans, Tilia, Vaccinium, Vigna sinensis, and Zanthoxylum clava-herculis. Oertel (1967, p. 10) helps account for the absence of Gossypium pollen in his statement, "Plant breeders have introduced a nectarless cotton so that destructive insects will not be attracted to the plant". Honey pollen analysis results concerning botanical origin did not consistently agree with beekeepers' reports. For example, sample 71 was reported as palmetto (Sabal minor) honey; this sample contained no Sabal minor pollen but had Rubus as dominant pollen and Trifolium repens as a secondary pollen. Louisiana honeys had fewer pollen grains per 10 g sample (Fig.l) than foreignproduced honeys. Over 50% of the Louisiana honeys had less than 20,000 grains per 10 g sample, whereas European workers (Maurizio, 1951) generally report that most honeys cleanly extracted by rotary extractor contain 20,000-100,000 grains per 10 g sample. European workers report that pollen content is increased in pressed or run honeys and is decreased when honey is adulterated with artificial products or by feeding sugar syrup to bees. In addition, the morphological structure of plants producing honey could affect the total pollen grain content. Louisiana beekeepers possibly produce less pollen-contaminated honeys than do European beekeepers, by using more refined techniques in their beekeeping and more efficient and cleaner extracting methods. Louisiana beekeepers may feed their bees more frequently, and thereby produce in some instances adulterated honeys with resulting lower pollen content. Oertel (1955) reported that nearly every beekeeper at some time or other feeds one or more colonies of bees.
122
MEREDITH HOAG LIEUX
The results presented in Table I indicate that the unifloral honeys of Trifolium repens, Berchemia scandens, and Rubus are to a considerable extent poor in pollen (< 20,000 pollen grains per 10 g). The Salix honey samples are for the most part richer in pollen (20,000-100,000 grains). No corrections were made to reestablish the probable nectar proportion associated with each pollen type as did Maurizio (1955, 1958) and Demianowicz (1964). Maurizio (1958) found most unifloral Trifolium repens samples in the 20,000-100,000 grain range. She .also found that three of five Salix samples had 20,000-100,000 grains; the other two Salix samples had over 100,000 pollen grains. Demianowicz (1964) reported that samples of unifloral Trifolium repens honey (samples obtained under controlled conditions) contained < 20,000 grains per 10 g sample. He also found that samples of Rubus honey were in the 20,000-100,000 grain range. A unifloral sample of Compositae honey (not shown in Table I) had the highest number of pollen grains per 10 g of any sample studied (approximately 3,300,000). This sample was probably polluted with pollen from pollen cells during the extracting process. Demianowicz (1964) reported that various Compositae (Asteraceae) honeys had total pollen grain counts per 10 g samples ranging from 1,950-43,480. A unifloral sample ofSapium sebiferum honey (not shown in Table I) had approximately 2,850 pollen grains per 10 g sample. The relatively large size of the Sapium sebiferum grain (33-36 # in diameter) possibly explains the low pollen count. Demianowicz (1964) concludes that there is an apparent relationship between the richness in pollen and the size of the grains: as a general rule, the smaller the pollen, the richer the honey. Louisiana honeys do not appear to have a particularly diverse botanical origin. Approximately 37% of the 54 Louisiana honey samples originated from six to ten different plant types. When combined, 75.9% of the samples originated from six to fifteen different plant types. Most of the Louisiana honey samples studied fell into the lower limits of Louveaux and Vergeron's 1964 study as to the pollen type ranges of Spanish honeys. For example, only 10% of Louisiana samples were in the 21-25 pollen type range.
Geographical origin Honeys collected from different honey producing areas of Louisiana are not easily distinguished from one another geographically by pollen types. Area 1 occupies chiefly the Mississippi River floodplain and delta and the floodplain of the Red River. The pollen types from honey gathered here indicate a varied flora of native and cultivated plants. The pollen assemblage represents plants growing on alternating low and higher ground with varying drainage, along streams, in woods, in swampy areas, in pastureland, and in cultivated fields. More honey is produced in this region than in any other area of the state. Three pollen types characteristic of Area 1 are: Cephalanthus occidentalis which indicates a swampy region, Mimosa strigillosa, and Forestiera
A MELISSOPALYNOLOGICALSTUDY OF 54 LOUISIANA (U.S.A.) HONEYS
123
acuminata which according to Brown (1945) is widely distributed in the Mississippi floodplain and along small streams and swamps. Maurizio and Louveaux (1965) reported that Mimosa pollen if found in honey could be used to distinguish exotic imported honeys fro_m central European honeys. Only one honey sample was collected from the pine-oak-hickory uplands of Area 2. Its pollen spectrum indicated no outstanding differences from the pollen assemblage of Area 1. This honey sample may be from an apiary located in a fringe area. Repeated attempts to collect more samples from area 2 were unsuccessful. Beekeepers living in this area reported one of two things: that they kept their bees in some other area or that the insecticide sprays used on cotton killed the bees and thus made commercial beekeeping unprofitable. Area 3 consists of pine-oak-hickory uplands and longleaf pine flatwoods. All samples collected in this area contained a characteristic pollen combination of Castanea and flex. There were no characteristic pollen types noted in either the prairie of area 4 or the pine-oak-hickory uplands and longleaf pine flatwoods of area 5. Area 6, the coastal marsh, is a low, wet plain along the Gulf of Mexico. For the most part, it is not suitable for beekeeping. Unifloral versus mixed honeys Of the 54 samples 31 were unifloral honeys (nine Trifolium repens, nine Rubus, seven Berchemia scandens, four Salix, one Sapium sebiferum, and one Compositae). All of the area 5 samples were unifloral honeys. There are probably more unifloral honeys produced in Louisiana than mixed honeys.
Floral versus honeydew honeys Most of the samples examined were clear colored and from nectar of flowers rather than from honeydew. But sample 40 (reported as a summer or fall honey; not included in the 54 samples from which data was recorded) may have originated, at least in part, from honeydew. Numerous fungus spores were observed. Zander (1949), as well as others, has reported that fungus spores are characteristic of honeydew honeys. Castanea pollen appeared as the most abundant pollen in sample 40. Barbier (1958) reported that Castanea produced both floral and honeydew honey that is rich in pollen. But Louisiana summer honeys often take up water and become dilute enough to ferment (K. Tucker, personal communication, 1971). Such conditions could account for sample 40 containing yeasts and fungus spores without the sample having necessarily derived from honeydew.
124
MEREDITH HOAG LIEUX
Season o f production Most samples examined were spring-produced honeys as indicated by their spring pollen spectra; a few were mixed spring and summer (e.g. samples 14, 26, 27, 38, 41, 76, 106, and 107); and possibly one sample was mainly summer or fall honey (sample 112). The pollen types used to identify late spring, summer, or fall honeys include: Glycine max, Cassia fasciculata, Polygonum, Lagerstroemia indica, L ythrum, Ampelopsis, Trachelospermum difforme, and others. The commercial beekeepers o f Louisiana probably sell mainly spring honey.
ACKNOWLEDGEMENTS I wish to recognize Dr. Clair A. Brown for his work on the photographs used in this manuscript and for the reference pollen he made available to me. I want to thank Dr. Shirley Tucker and Dr. Abdel Wahab. I. Allam (A. I. Allam) for reading the manuscript and giving technical advice. Due acknowledgement is made to National Science Foundation (U.S.A.) for their financial support during this study.
REFERENCES Barbier, E., 1958. Examen pollinique de quelques miels unifloraux. Ann. Abeille, 1: 73-76. Brown, C. A., 1945. Louisiana Trees and Shrubs. Louisiana For. Comm. Bull., 1:262 pp. Demianowicz, Z., 1964. Characteristik der Einartenhonige. Ann. Abeille, 7: 273-288. Erdtman, G., 1952. Pollen Morphology and Plant Taxonomy. (An Introduction to Palynology, 1 Angiosperms). Almquist and Wiksell, Stockholm, 539 pp. Louveaux, J. and Vergeron, Ph., 1964. Etude du spectre pollinique de quelques miels espangnols. Ann. Abeille, 7: 329-347. Maurizio, A., 1951. Pollen analysis of honey. Bee World, 32: 1-5. Maurizio, A., 1955. Beitr~igezur quantitativen PoUenanalyse des Honigs. Z. Bienenforsch, 3: 32-39. Maurizio, A., 1958. Beitr~igezur quantitativen PoUenanalyse des Honigs. Ann. Abeille, 1: 93-106. Maurizio, A. and Louveaux, J. 1965. Pollens de plantes Melif~res d'Europe. Union des Groupements Apicoles Fran~ais, Pads, 148 pp. Oertel, E., 1939. Honey and Pollen Plants of the United States. U.S. Dept. Agricult. Circ., 544, 63 pp. Oertel, E., 1955. The Beginner Beekeeper in Louisiana. Louisiana State Dept. Agricult. Immigr. Baton Rouge, La., 3rd ed., 46 pp. Oertel, E., 1967. Nectar and pollen plants. U.S. Dept. Agricult. Handbook, 335: 10-16. The Louisiana State Department of Agriculture, 1966. Louisiana-A Few Concrete Facts. Louisiana State Dept. Agriculture, Baton Rouge, La., 75 pp. Zander, E., 1935. Beitr~igezur Herkunftsbestimmung bei Honig. Pollengestaltung und Herkunftsbestimmung bei Bliitenhonig. Reichsfachgruppe linker, Berlin, 1:343 pp. Zander, E., 1949. Beitrage zur Herkunftsbestimmung bei Honig. Studien zur Herkun]tsbestimmung bei Waldhonigen. Franz Ehrenwirth, Miinchen, 4:267 pp.
A MELISSOPALYNOLOGICAL STUDY OF 54 LOUISIANA (U.S.A.) HONEYS
MEREDITH HOAG LIEUX 1
Department o f Botany, Louisiana State University and A. and M. College, Baton Rouge, La. (U.S.A.) (Received August 6, 1970) (Resubmitted September 1, 1971)
ABSTRACT Lieux, M. H., 1972. A melissopalynological study of 54 Louisiana (U.S.A.) honeys. Rev. Palaeobot. Palynol., 13: 95-124.
Pollen analyses were made of 54 commercial Louisiana (U.S.A.) honeys collected during 1967-1968. Fifty-eight different pollen types were identified. A direct correlation was assumed between a plant's pollen quantity and its nectar contribution. On this basis, the major Louisiana honey plants that were determined are: Fabaceae, predominantly Trifolium repens (99%), Rubus, Berchemia seandens, and Salix. Plants established as important for their nectar contributions in isolated samples axe: Cephalanthus occidentalis, Sapium sebiferum, miscellaneous Compositae (Asteraceae), Glycine max, and Parthenocissus quinquefolia. Native plants such as Rubus, Berchemia scandens, and Salix contribute more to Louisiana's honey sources than do cultivated plants or introduced plants. Most samples originated from six to fifteen different plant types and thus do not appear to be of particularly diverse botanical origin. The pollen types that indicate late spring, summer, or fall honeys include: Glycine max, Cassiafasciculata, Polygonum, Lagerstroemia indica, Lythrum, Ampelopsis, Trachelospermum difforme, and others. Thirty-one samples could be considered unifloral honeys. Most Louisiana honeys were a clear color and from nectar of flowers and not honeydew. Honeys collected from different regions of Louisiana are not easily distinguished from one another geographically by pollen types except in a few instances. Three pollen types characteristic of Mississippi and Red River floodplain honeys are: Cephalanthus occidentalis, Mimosa strigillosa, and Forestiera aeuminat~ Castanea and llex form a characteristic pollen combination in honeys produced from the pine-oak-hickory uplands and longleaf pine flatwoods of western Louisiana.
1 Present address: Department of Geology, Louisiana State University and A. and M. College, Baton Rouge, La. 70803 (U.S.A.)
96
MEREDITH HOAG LIEUX
INTRODUCTION Microscopical analysis of honey is based on the fact that the raw materials of honey (nectar and honeydew) have certain constituents which remain identifiable in the ripe honey. For the nectar, these are mainly pollen grains from the blossoms producing the nectar. For the honeydew, these constituents are usually algal cells, fungus spores, and fungus hyphae that come from the surface plants of forest trees. Other materials present in honey include living organisms (especially yeasts), insect body parts, soot, and dust particles. The chief objectives of microscopical examination of honey are to assess the value of different plant species as nectar sources; to determine the geographical origin of honey through its pollen spectrum; to estimate the honeydew contributions by means of fungus spores, fungus hyphae, and algae; to diagnose the poisoning of bees from the poisonous plants they visit; to ascertain the season of production; to check for pollution and adulteration of honey. Results obtained from microscopical analysis of honey concern beekeepers, the honey trade, and State Food Control offices. Beekeepers are especially interested in determining the botanical origin of honey. They want to locate their hives so as to fully utilize existing honey sources. The honey trade seeks knowledge concerning the botanical and geographical origin of honey to help in establishing quality standards. State Food Control offices in many countries require that the quality of the product stated on the label conforms to reality. Furthermore, some countries with high production costs legally protect domestic honey. They enforce a compulsory declaration of origin which can only be determined by microscopical analysis of honey. In several European countries (e.g. France, Germany, Sweden, and Switzerland) honey analysis research is progressing rapidly towards locating and defining the native honey types. But in the United States there is need for comprehensive inquiry on the subject. This research is a melissopalynological investigation of 54 commercial Louisiana (U.S.A.) honeys in which the pollen grains of entomophilous plants predominate. Its purpose is to study the honeys in regard to botanical origin, geographical origin, season of production, unifloral versus mixed honey, and floral honey versus honeydew honey. This work is the first major pollen study of Louisiana honeys.
METHODS
Collection and preparation of reference pollen A reference slide collection of Louisiana honey pollen was prepared. Pollen samples and herbarium specimens were collected from many plants; herbarium specimens were identified; pollen samples were processed and mounted. The pollen samples were prepared by a slight modification of the Erdtman (1952) acetolysis technique
A MELISSOPALYNOLOGICAL STUDY OF 54 LOUISIANA (U.S.A.) HONEYS
97
(chlorination was omitted). Brief identification notes pertaining either to the honey plants or their pollen were entered on cards and assembled in a reference card f'de.
Collection and preparation of honey samples A 1967 list of commercial beekeepers in Louisiana was obtained from the Division of Entomology, Louisiana Department of Agriculture and Immigration. Selected commercial beekeepers throughout the state of Louisiana were contacted and requested to collect uncontaminated (not mixed with samples from other locations) commercial type honey samples in their given areas during the fall of 1967; spring, summer, and fall of 1968. Pollen was extracted from a 20 g honey sample by centrifugation following dilution (20 cm 3 water) and heating (45 °C). The recovered pollen sediment was treated according to the acetolysis schedule. Enough glycerin jelly was added to give the acetolyzed honey pollen sediment a 1 cm 3 volume. A permanent reference slide was made of each sample. Temporary slides for counting purposes were made using the following technique: (1) A vial containing the pollen sediment-glycerin jelly mixture was heated until the glycerin jelly was quite fluid. (2) The sample was stirred with a warm glass stirring rod. (3) An eyedropper was warmed and used to place one drop of sample on a warm clean slide. The same eyedropper, thoroughly cleaned after each use, was used to prepare every temporary counting slide. (4) A 22 mm 2 (no. 1 or 0) cover slip was centered over the drop. Care was taken to assure that the sample covered the 22 mm 2 area evenly and that none leaked from the sides. (Gentle warming of the slide helped in spreading the drop).
Examination of honey samples and data collection The species frequency percentages were based upon a count of at least 250 pollen grains per sample. Up to eleven traverses per slide (at predetermined points 2 mm apart) were made under high power (40×). The first 250 grains were identified and counted. A traverse was completed even though a 250 grain count was reached beforehand. Thus, in most instances more than 250 grains were recorded. If grains were too scarce on the first temporary slide, two or more slides were made and counted until the 250 mark was reached. The 1 cm 3 sediment-glycerin jelly mixtures that had grains too abundant to allow accurate counting were diluted with 1-3 cm 3 of glycerin jelly before making the temporary slides. An approximate number of pollen grains per 20 g honey sample was calculated as follows (this number was halved for comparative purposes and represents a 10 g sample in the results): N = total number of grains per 20 g honey sample; n = number of grains counted; A = total area of 22 mm 2 cover slip (484 mm2); a = area covered by
98
MEREDITH HOAG LIEUX
11 traverses (108.9 mm2), diameter of microscopic field was 0.45 mm; k = area adjustment constant (4.3); ?~ = (1/a)A; d = number of drops per cc (24 drops). The formula (N = ?~nd) was used in calculating total grain count of the original 20 g honey sample reduced to 1 cm 3. Adjustments to the formula were necessary, if fewer or more than eleven traverses were made, and if the 1 cm 3 sample was diluted with more glycerin jelly. The classification of Zander (1935) was used in this study to indicate pollen and associated nectar source: dominant pollen (over 45%), secondary pollen (16-45%), minor pollen (1-15%); trace pollen (less than 1%) was added for convenience. A dominant pollen (over 45%) denotes a unifloral honey. A direct correlation was assumed between a plant's pollen quantity and its nectar contribution to the honey. RESULTS Total grain counts were made on each of the 54 honey samples. With this data, the number of samples appearing within certain arbitrarily set ranges of pollen grain totals was determined (Fig.l). Of the 54 honey samples studied, 28 (51.9%) had less than 20,000 grains per 10 g; 12 (22.2%) had 20,000-100,000 grains; 14 (25.9%) had more than 100,000 grains. The unifloral honey samples of Trifolium repens I , Rubus, Berchemia scandens, and Salix were arranged in a Maurizio-type table (Maurizio, 1958, p. 105). The Trifolium repens and the Salix type samples fell into two discrete ranges, while the Rubus and Berchemia types fell into three continuous ranges (Table I). The first three unifloral type
TABLE I Pollen analyses of 29 unifloral 10 g Louisiana honey samples collected through 1967-1968 Dominant pollen types
Percentage
Number of honey samples within the indicated range of amount of pollen grains 20,000 20,000-100,000 2> 100,000
Total number of honey samples
Trifolium repens I Rubus Berchemia scandens Salix
45-92 51-93 49-83 45-53
6 5 4 1
9 9 7 4
2 1 3
3 2 2 -
1Fabaceae, predominantly Trifolium repens (99%). samples (Table I) showed pollen grain number in the lower and higher limits of the selected range. Salix unifloral type did not contain any 2> 100,000 pollen grain samples. 1Trifolium repens will refer throughout the text to Fabaceae, predominantly (99%) T. repens.
A MELISSOPALYNOLOGICALSTUDY OF 54 LOUISIANA (U.S.A.) HONEYS
99
29
E
E
2
14 12
( 20,000
20,000-100,000
7100, O00
Pollen grains in 10g honey samples Fig.1. Frequency of honey samples in the range ~ 20,000; (20,000-100,000); ~ 100,000 total pollen grains per 10 g sample.
Respectively 66%, 55%, and 25% of Trifolium repens, Rubus, Berchemia scandens, and Salix samples showed < 20,000 pollen grains in 10 g of honey. Fifty-eight pollen types were identified in the 54 honey samples (Fig.2). (Photographs of most of the 58 pollen types are included in Hate I-VIII). The pollen grains were determined to the family, genus, or species (when possible). Percentage frequencies of the pollen types were calculated and shown in a bar graph (Fig.2). Eight pollen types appeared in 50% or more of the honey samples. In descending order, they are Berchemia scandens, Rubus, Trifolium repens, Salix, Rhus radicans, Rosaceae, Quercus, and Gleditsia. Thirty pollen types appeared in 10% or less of the samples. Fig.3, a Louveaux and Vergeron-type graph (Louveaux and Vergeron, 1964, p. 340), shows a straight-line trend and gives a true picture of Louisiana honey origin. The number of different pollen types per sample is shown (Fig. 4). Of the 54 samples studied, 75.9% had six to fifteen pollen types; 22.2% had sixteen to twenty-five pollen types; and 1.9% had one to five pollen types.
MEREDITH HOAG LIEUX
100
Pollen types
0
IO
20
30
60 50 40 Percent frequency
70
80
Fig.2. Percent frequency of 58 pollen types in 54 Louisiana honey samples.
Pollen spectra by percentages for the 54 samples were grouped according to modifications of Oertel’s (1955) beekeeping areas that are outlined in Fig.5. (These areas were determined on the basis of geology, soil, topography, and vegetation differences). The pollen spectra by percentages are given in Table U-VI. Composite pollen spectra for areas 1, 2,3,4, 5, and for the state are shown in Table VII.
90
100
A MELISSOPALYNOLOGICAL STUDY OF 54 LOUISIANA (U.S.A.) HONEYS
58
55 ,r. ~. z. :~ = _ f . - . . . .
50
~
45
g ~. 4o ~ 35 E ~, 30
Number of honey samples (54)
Fig.3. Straight-line trend representing 54 Louisiana honey samples as to the 58 identified pollen types.
Q_
E
x:l
E
3
1
I
3 (I-5)
8 (6- I0)
13 (11= 15)
18 (16-20)
23 +- P.- Type Average (21-25) (- P.- Type Range
Number of pollen types (58)
Fig.4. Histogram showing range of different pollen types within honey samples.
101
MEREDITH HOAG LIEUX
102
PLATEI
I
0
E
G
I( |
!
"50u
Pollen grains representing pollen types identified in the boney samples studied. A. Allium canadense L. (106-104A). B,C. Ampelopsis arborea (L.) Koehne (160-160A). D,E. Berchemia scandens (Hill) K. Koch (111-109A). F,G. Brassica (31-24A). H. Brunnichia cirrhosa Gaertn. (166-169A). J,K. Callicarpa americana L. (25683-238A). Magnification X 900. Code number refers to the herbarium specimen and the reference pollen slide in the Lieux collection.
A MELISSOPALYNOLOGICAL STUDY OF 54 LOUISIANA (U.S.A.) HONEYS
103
P L A T E II
Oi) I
L
•
!
,50u
Pollen grains representing pollen types identified in the honey samples studied. A,B. Cassiafasciculata Michx. (188-192A). C,D. Castaneapumila (L.) Mill. (404064-236A). E,F. Celtislaevigata Willd. (73-87A). G,H. Cephalanthus occidentalis L. (158-157A). J. Cocculuscarolinus (L.) DC. (Brown 1261). K,L. Cornusdrummondi Meyer (Brown 775). M,N. Cornusflorida L. (99-59A). Magnification X 900. Code number refers to the herbarium specimen and the reference pollen slide in the Lieux pollen collection.
104
MEREDITIt HOAG LIEUX
P L A T E III
4
8
I |
,( it ,
,,,
,50U
Pollen grains representing pollen types identified in the honey samples studied. A,B. Forestiera acuminata (Michx.) Poir. (462328-232A). C,D. Fraxinusamericana L. (171488-233A). E,F. Gleditsia aquatica Marsh. (Brown 1/61). G,H. Gleditsia triacanthos L. (137723-227A). J,K. Glycine m a x (L.) Merr. (196-194A). L,M llex cornuta Lindl. (57-55A). Magnification X 900. Code number refers to the herbarium specimen and the reference pollen slide in the Lieux pollen collection.
A MELISSOPALYNOLOGICAL
STUDY
OF 54 LOUISIANA
(U.S.A.)
JIONEYS
105
PLATE IV
I
Pollen A. B,C. D. E. F. G. H. J,K. L,M.
ISOU
I
grains representing pollen types identified in the honey samples Lagerstroemia indica L. (Brown 1585). Ligustrum (154-152A). Liquidambar styraciflua L. (46A). Lythrum alatum Pursh. (190-190A). X 450. Magnolia grandiflora L. (154-153A); magnification Mikanea scandens (L.) Willd. (Brown 1375-18761). Mimosa strigillosa Torrey et Gray (186-165A). Nyssa sylvatica Marsh. var. biflora (Walt.) Sarg. (Brown 746). Parthenocissus ipinquefolia (L.) Planch. (335458-219A).
Magnification X 900, unless otherwise stated. Code number reference pollen slide in the Lieux pollen collection.
studied.
refers to the herbarium
specimen
and the
106
MEREDITH HOAG LIEUX
PLATE V
,,L
C E
IP
U
It
1
i
,50u
Pollen grains representing pollen types identified in the honey samples studied. A. Planera aquatica (Walt.) J.F. Gmel. (99318-231A). B. Polygonum (Brown 1226-18541). C,D ,E.Prunus caroliniana Mill. (27-20A). F,G. Prunuspersica (L.) Batsch (33-23A). H. Prunus serotina Ehrh. (63-37A). J. Pyrus (20-15A). Magnification X 900. Code number refers to the herbarium specimen and the reference pollen slide in the Lieux pollen collection.
107
A MELISSOPALYNOLOGICAL STUDY OF 54 LOUISIANA (U.S.A.) ItONEYS PLATE VI
B
E
i,
F
G
L
K tt
M
V
!
I
,50u
Pollen grains representing pollen types identified in the honey samples studied. A. Quercus rnichauxii Nutt. (Brown 774). B,C,D. Rhus radicans L. (Brown 963-17728). E,F,G. Rubus (40-36A). Rumex crispus L. (Brown 759). H,J. K,L,M. Salix nigra Marsh. (23-16A). N,O,P. Sapium sebiferum (L.) Roxb. (Brown 1584). Magnification X 900. Code number refers to the herbarium specimen and the reference pollen slide in the Lieux pollen collection.
108
MEREDITH HOAG LIEUX
PLATE VII
A
l
E V
,d•
H
wll
,,
,
,
.......
, 50U
Pollen grains representing pollen types identified in the honey samples studied. A,B. Senecio glabellus Poir. (44-44A). C. Trachelospermumdifforme (Walt,) Gray (446838-206A). D,E. Trifoliurn incarnatum L. (65-79A). F,G. Trifolium repens L. (86-73A). H. Ulmusfulva Michx. (Brown 762). J,K. Verbenarigida (L.) Spreng. (Brown 1244-18587). Magnification X 900. Code number refers to the herbarium specimen and the reference pollen slide in the Lieux pollen collection.
A MELISSOPALYNOLOGICAL STUDY OF 54 LOUISIANA (U.S.A.) HONEYS
109
PLATE VIII
r.
A
!1
t,,,
Pollen A,B. C,D. E,F.
D
I
'50u
grains representing pollen types identified in the honey samples studied. Vicia ludoviciana Nutt. (95-63A). Vitis (Brown 729). Zea mays L. (Brown 1290);magnification X 450.
Magnification X 900, unless otherwise stated. Code number refers to the herbarium specimen and the reference pollen slide in the Lieux pollen collection.
I
• 106
60
Fig. 5. Beekeeping areas of Louisiana.
S C A LE
LOUISIANA
55 "56
9= ~.git.4,
we.t
.I
/
I
C ~ n~
k 91
Pine-Oak-Hickory Pine-Oak-Hickory Uplands and Longleaf Pine Flatwoods Prairie Longleaf Pine Flatwoods and Upland Hardwoods Marsh Lands
Area Q Area Q Area Q Area O Area G
i
*Small numbers refer to sampling sites.
Uplands
Bottomland Hardwoods and Cypress
Beekeeping Areas of Louisiana (USA)
Area O
Figure 5.
c
© > o
~D
A MELISSOPALYNOLOGICAL STUDY OF 54 LOUISIANA (U.S.A.) HONEYS
111
TABLE II Percentages of pollen types for honey samples from area 1
Taxa~ples
Acer Allium Ampelopsis Berchemia scandens Brassica Brunnichia cirrhosa Cassia fasciculata Castanea Celtis Cephalanthus occidentalis Cercis canadensis Chenopodium Cocculus carolinus Compositae Comus Cruciferae Diospyros virginiana Forestiera acuminata Fraxinus Gleditsia Glycine max Gramineae llex Leguminosae (Trifolium repens) L igu s tru m Liquidambar styraciflua Lythmm Mimosa strigillosa Nymphaea Nyssa Parthenocissus Pinus Planera aquatica Polygonum Quercus Rhus radicans Rosaceae Rubus Rumex Sagittaria Salix Sapium sebiferum Scrophulariaceae I (?) Trachelospermum difforme l Tentative identification.
10
11
14
17
18
26
31
27
0.4 0.8
12.1
0.9
28.5
1.1
5.9
0.4 2.6
6.7
0.9 0.4 29.0
3.1
1.0
0.4 0.4
0.4
1.7
0.4
0.4 3.1
0.4
0.4
0.4 0.4 0.4 0.8 9.0
66.3
1.6
1.1
6.7
0.4 4.7
0.4 1.5
1.8
2.2 0.4 42.1
1.2 7.8 3.9 7.0
1.1 1.1 3.4 1.1 4.1
9.0
48.9
39.6
18.4 0.9 16.0 0.4
0.7
1.1 2.7 1.1 38.9
41.2
0.4 10.3 3.9 21.3 0.4 O.7 30.9
0.9
50.9
3.0
43.8
7.9
0.9
0.4 1.1
0.4 0.7 44.0 0.4 0.4 10.8
0.7
12.6
0.4 1.5
0.4
17.5 1.1
0.4
1.9 3.7 1.1
4.1
MEREDITH HOAG LIEUX
112 TABLE 1I (continued)
•
Samples
10
11
14
17
18
26
27
0.4
2.7
1.1
31
Taxa
Trifolium incarnatum Ulmus Umbelliferae
Verbena Vicia Vitis
1.1
0.9
1.7
0.4 0.7
0.4 0.4
2.4 2.4 0.4 1.2 0.8
44
46
51
62
63
64
1.7
6.4
1.6
20.3
1.4 10.4
4.2 6.5
0.7
UK-11 UK (Unknown) UI (Unidentifiable) mples
A cer Allium Ampelopsis Berehemia seandens Brassiea Bnmniehia cirrhosa Cassia faseiculata Castanea Celtis Cephalanthus occidentalis Cercis canadensis Chenopodium Cocculus carolinus
0.4 0.8
0.4
33
43
0.4
5.2 0.4
1.5 1.1
0.4 1.9
1.9
1.9
3.8 0.6
0.4
Compositae
0.4
Comus
0.4
1.2 0.4
0.3 1.5
1.1 0.7
0.9
0.4
4.9
1.8 6.8
0.9
0.7
42.1
37.4
Cruciferae
Diospyros virginiana Forestiera acuminata Fraxinus Gleditsia Glycine max Gramineae
3.8
0.5
3.8
1.2
flex Leguminosae (Trifolium repens) Ligustrum Liquidambar styraciflua Lythrum Mimosa strigillosa Nymphaea Nyssa Parthenoeissus Pinus
15.0
82.1
13.8
40.6
66.0 0.4
21.3
0.4 0.8
6.2
1.5 0.3
0.4 0.7
0.6
113
A MELISSOPALYNOLOGICAL STUDY OF 54 LOUISIANA (U.S.A.) HONEYS TABLE II (continued)
•
Samples
33
43
44
46
51
62
63
64
0.9 0.5 75.1
8.0 0.4 32.2
3.0 0.8 9.0
0.3 4.0 0.6 0.9
6.1 1.8 3.6
7.4
5.9 0.2
10.0
5.1
15.2
24.1
53.4
Taxa
Planera aquatica Polygonum Quercus Rhus radicans Rosaceae Rubus Rumex Sagittaria Salix Sapium sebiferum Scrophulariaceae (?) Trachelospermum difforme Trifolium incarnatum Ulmus Umbelliferae Verbena Vicia Vitis UK-11 UK UI ~ Samples Taxa " ~
Acer Alliurn A mpelopsis Berchemia scandens Brassica Brunnichia cirrhosa Cassia fasciculata Castanea Celtis Cephalanthus occidentalis Cercis canadensis Chenopodium Cocculus carolinus Compositae Comus Cruciferae Diospyros virginiana Forestiera acurninata Fraxinus
0.4 8.6 34.0 0.8
15.0
3.5
3.8
1.5
0.3
1.9 0.4
0.4
1.1
0.4
1.6 9.4
2.1
0.9 0.3
1.9 0.4 3.8 1.9 65
0.8 1.9 66
0.8 1.6
1.4 67
68
71
2.1 0.3 1.8 72
2.2 73
1.8 75
0.5
42.6
8.6 O.8
52.7
26.3
6.9
39.2
10.0
0.6
0.2 0.9
O.2 5.8
0.7 0.3
0.3
0.3
0.4
2.5 0.5
0.4
1.0
0.3
0.5 0.5
0.2
O.2 0.3 0.3
114
MEREDITH HOAG LIEUX
TABLE II (continued)
Taxa~amples
65
66
67
68
Gleditsia Glycine max Gramineae llex Leguminosae (Trifolium repens) Ligustrum Liquidambar styraciflua L ythrum Mimosa strigillosa Nymphaea Nyssa Parthenocissus Pinus Planera aquatica Polygonum Quercus Rhus radicans Rosaceae Rubus Rumex Sagi t taria Salix Sapium sebiferum Scrophulariaceae(? ) Trachelospermum difforme Trifolium incarnatum Ulmus Umbelliferae Verbena Vicia Vitis UK-11 UK UI
3.6
5.8
2.0
0.7
15.5
0.3 1.0
0.4 0.4 43.6
~
les
Acer Allium Ampelopsis Berchemia scandens Brassica Brunnichia cirrhosa Cassia fasciculata Castanea Celtis
0.3 16.7
71
72
73
75
0.3
0.7
2.5
0.2 23.5
14.9
54.8
0.2 45.4
0.2 11.6
0.9 1.2
0.2
0.8
0.4
0.9 1.8 3.2 ",.-,~
19.0 0.3
1.9 9.3 1.2 35.4
0.5 3.0 8.5 13.4
0.4 15.2
13.4
0.3
0.6 O.3 0.9 0.3 0.3
0.3 0.9 26.3
16.9
5.4
0.5 7.3 0.9 2.7 0.2
3.7 1.3 7.8
17.9
18.2 0.2
0.7
0.2
0.2 0.2 0.2 0.5
1.0
0.4 0.4
0.8
76
79
85
67.4
3.7
27.7
0.3
3.8 2.1 59.2
0.3
0.8 2.3
0.7
3.3 0.7 16.9 0.4
1.5 86
33.2
0.4 1.4
0.3
87
93
107
11.4
12.8
14.4
7.6 0.4
0.2
0.4 0.5 0.3 0.3 0.2
A MELISSOPALYNOLOGICAL STUDY OF 54 LOUISIANA (U.S.A.) HONEYS
115
TABLE II (continued)
Samples
76
79
85
86
87
93
107
Taxa
Cephalanthus occidentalis Cercis canadensis Chenopodium Coeeulus earolinus Compositae
0.3
1.9
0.3 0.4 0.7
Cornus
0.3 0.3
0.4
0.3
6.1
Cruciferae
Diospyros virginiana Forestiera acuminata Fraxinus Gleditsia Glycine max
1.0 0.7
0.8 0.3 0.5
Gramineae
Ilex Leguminosae (Trifolium repens)
Ligustrum Liquidambar styraeiflu Lythrum Mimosa strigillosa Nymphaea Nyssa Parthenocissus Pinus Planera aquatiea Polygon u m Quercus Rhus radieans
13.3
0.4 0.4 6.8
0.3 18.7 0.3
0.4 0.4
6.0
5.9
0.3
0.3 0.2 11.8
1.7
5.3
0.6 1.0
1.3
0.3 6.7
7.6
1.5
0.3 O.8
Rosaceae
Rubus Rumex Sagittaria Salix Sapium sebiferum
56.4
7.9 2.3 1.2
0.6 0.6 1.8 0.3 3.8 2.1 0.9
O.3 O.3
0.3 4.8 2.7 2.7
0.4 2.4 5.2 35.4
0.6 7.6 2.0 28.3
0.8 3.0 1.1 13.6
0.4 8.1 3.8 41.5
2.6 3.8 17.4 19.8
11.0 0.3
17.1 0.8
1.7 1.2
45.4
24.3 0.4
18.4
Serophulariaceae (?)
Traehelospermum difforme Trifolium inearnatum Ulmus
2.3
Umbelliferae
0.5
Verbena Vieia Vitis
O.3
UK-11 UK U!
O.3
1.3
0.3 0.3 0.3 0.5
0.3
0.4
0.3
0.8 0.4 0.8
0.2
0.3 0.3
0.6
0.9
116
MEREDITH HOAG LIEUX
TABLE 1II Percentages of pollen types for honey samples from area 2
Taxa
Sample 30
A Ilium 1.0 Berchemia scandens 41.1 Brassica 0.4 Castanea 3.1 Cephalanthus occidentalis 3.5 Comus 0.4 Diospyros virginiana 0.7 Leguminosae (Trifolium repens) 13.2 Ligustrum 0.4 Magnolia 1.0 Mimosa strigillosa 0.7
Taxa
Sample 30
Parthenocissus Quercus Rhus radicans Rosaceae Rubus Salix Sapium sebiferum Verbena Vitis UI
13.2 0.4 2.4 2.1 3.5 4.5 1.4 1.4 4.2 1.4
TABLE IV Percentages of pollen types for honey samples from area 3
Taxa~
arnples
Berehemia seandens Brassica Brunniehia cirrhosa Castanea Celtis Compositae Comus Diospyros virginiana Fraxinus Gleditsia Gramineae Ilex Lagerstroemia indica Leguminosae (Trifolium repens) Ligustrum Liriodendron tulipifera Magnolia Myrica Nyssa Parthenocissus Pinus
41
55
56
57
60
61
84
29.4
26.6
37.9 O.4
49.8
81.7
28.0
56.3
4.5 6.0
10.0
9.2
5.5
4.0
1.7
4.4 0.4 0.4
0.4
0.4
1.0 0.4
0.4 0.4 1.1
1.9 3.0 6.0
3.5
0.4 1.0
0.7
4.4
0.7 3.8
3.5
4.5
0.3 0.4 0.4 0.4 3.4 17.0 0.4
0.3
1.5
14.4
0.4 0.4
2.8
1.4
2.6
117
A MELISSOPALYNOLOGICAL STUDY OF 54 LOUISIANA (U.S.A.) HONEYS TABLE IV (continued)
Samples
41
55
0.4 0.7 0.7 2.3 12.0 0.4 3.4
5.9 1.0 5.5
56
57
60
61
84
Taxa Polygonum Quercus Rhus radicans Rosaceae Rubus Sagittaria Salix Sambucus Sapium sebiferum Scrophulariaceae (?) Trifolium incarnatum Vitis Zea mays UK-11 UK UI
45.9
0.4 15.2 25.9 6.4 0.4
0.4 6.3 1.1 4.1
0.8
29.1
56.8
11.1
1.7
0.7 1.1
0.7 5.6 0.4 0.4 2.7 0.4 1.9
1.0
0.4
0.4
0.4
0.4 1.0
0.7
2.0
1.0
1.1
TABLE V Percentages of pollen types for honey samples from area 4
T~a~amples
35
Berchemia scandens 37.5 Cassia fasciculata Castanea Compositae 1.9 Comus Fraxinus Gleditsia 0.4 Glycine max Gramineae llex Leguminosae (Trifolium repens) 12.1 Ligustrum Magnolia Parthenocissus Quercus Rhus radicans 8.3 Rosaceae 11.7
38
59
69
0.7 2.9
8.6
8.7
5.0
0.6
1.2 0.8
0.3
29.4
2.8
42.3 0.8
89
20.8
106
6.3
0.5 0.3 0.8 2.0
0.4
40.5 0.4 46.0
70
0.8
0.3
91.6
20.2
5.0 3.1 0.5 2.7 0.9
0.7 1.2
0.4 0.4 3.2 2.3
0.3 12.1 2.2
1.3 0.5
118
MEREDITH HOAG LIEUX
TABLE V (continued)
Samples
Rubus Salix Sapium sebiferum Trifoliurn incarnatum Ulmus Verbena Vitis Zea mays UK UI
35
38
59
69
70
89
106
15.9 9.5 0.4
0.4 0.4
50.7 8.1
20.1 3.2 13.4
3.6
25.6 9.0 2.5 1.4
6.8 2.2 65.6
0.4 0.4
0.7 0.4
0.1
1.1
0.7 0.4 1.5
3.2
0.2
0.8
0.9 0.5
0.3 1.4
1.4 1.8
100
102
103
49.3
83.4
16.3
0.8
TABLE VI Percentages of pollen types for honey samples from area 5
T a x a ~Samples
Berchemia scandens Callicarpa americana Chenopodium
37
2.7
97
98
21.1
27.8
99
2.8
0.3 2.1 86.6
Compositae
Comus Fraxinus
27.1
0.8
0.7 0,4
Gramineae
llex Leguminosae (THfolium repens) Ligustrum Magnolia Nyssa Parthenocissus Quercus Rhus radicans
0.4 1.5 0.4 0.8
0.4 2.2
0.7 0.7 0.3
1.4
4.8 0.4 0.4
1,1 3.3
2.2
0.4
0.4 0.3 2.5 0.1
0.4
1.2 0.8 3.1
4.2 1.9
60.1
73.5
1.5 0.4 61.5
1.1
0.4
3.3
Rosaceae
Rubus Sagittaria Salix Sapium sebiferum
112
0.4 0.3 92.8
19.7
6.6
11,3 8.5
0.5 76.7
3.8 0.1
0.7
Scrophulariaceae (?)
0.5
Trifolium incarnatum Vicia Vitis
2.7
UK UI
0.4 0.8
0.4 1.3 0.7 0.4
1.1
1.4 0.7 0.3
2.8 2.8
0.7 1.1
0.4
0.1 0.2
A MELISSOPALYNOLOGICAL STUDY OF 54 LOUISIANA (U.S.A.) HONEYS
119
TABLE VII Composite pollen spectra by percentages for honey samples from the five main honey producing areas of Louisiana, plus a composite state spectrum
Taxa
Area 1
Acer Allium Ampelopsis Berchemia scandens Brassica Brunnichia cirrhosa Callicarpa americana Cassia fasciculata Castanea Celtis Cephalanthus occidentalis Cercis canadensis Chenopodium Cocculus carolinus
0.01 0.07 0.2 15.2 0.02 0.02
Compositae
Comus Cruciferae
Diospyros virginiana Forestiera acuminata Fraxinus Gleditsia Glycine max Gramineae
Rosaceae
Area 3
Area 4
Area 5
41.1 0.4
44.2 0.05 0.6
0.4
0.2
0.8
12.2
25.4
0. 03
0.01
3.1
0.2 1.5 0.05 0.03 0.01 0.6 0.1 0.03 0.04 0.4 0.3
0.05 3.5 0.3
0.4
0.2 0.1
0.7
O.1
1.4 2.0 0.3 0.1
3.2 17.9
5.8
13.2 0.4
1.3 0.04
0.05
0.1
0.2
0.4 6.5 0.7 0.07
0.2 2.4 0.4 2.0 2.1
10.8 3.6
0.05
0.1 1.0
34.9 0.1
1.5 0.05
0.1
0.06
0.05 0.05
0.5 0.2
0.05 1.0 0.7
0.05
State
0.1 0.06 0.1 20.6 0.02 0.1 0.01 0.2
1.0
0.3
Ilex Lagerstroemia indica Leguminosae (Trifolium repens) 26.1 Ligustrum 0.02 Liquidambar styraciflua 0.02 Liriodendron tulipifera Lythrum 0.01 Magnolia Mimosa strigitlosa 1.6 Myrica Nymphaea 0.01 Nyssa 0.2 Parthenocissus 1.7 Pinus 0.01 Planera aquatica 0.03 Polygonum 0.04 Quercus 0.2 Rhus radicans 4.5 Rub'us
Area 12
0.05
0.1 0.7 0.03 0.06 0.01 2.1 0.6 0.02 0.06 0.2 0.2 0.9 2.0 0.3 0.5 0.06 20.3 0.3 0.01 0.01 0.01 0.05 1.0 0.01
0.01 13.2
0.4 2.4 2.1 3.5
1Area 2 is represented by only one honey sample.
1.6 2.5 0.05 0.05 0.2 4.0 0.6 16.6
0.3 3.5 2.5 17.6
0.6 0.9 0.08 49.3
0.4 1.6 0.1 0.02 0.03 0.5 3.7 2.3 22.1
MEREDITH HOAG LIEUX
120 TABLE VII (continued)
Taxa
Area 1
Rumex Sagittaria Salix Sambucus Sapium sebiferurn Scrophulariaceae (?) Trachelospermum difforme Trifolium incarnatum Ulmus Umbelliferae Verbena Vicia Vitis Zea mays UK-II UK UI
0.03 0.03 17.9
Area 2
4,5
O.1 0.1 0.01 0.09 0.3 0.01 O.2 0.8 0.3
1.4
4.2
0.2 0.4 1.0
1.4
Area 3
0.05 12.5 0.05 O.2 0.1 0.8
1.4
Area 4
Area 5
4.6
0.01 2.1
11.6
1.1 0.06
0.2 0.2
0.4
O.7 0.4 0.05 0.4 0.1 1.1
0.2 0.1 0.3 1.0
0.2 0.6 0.2 0.9
DISCUSSION This research presents information on Louisiana commercial honeys concerning botanical origin, geographical origin, unifloral versus mixed honey, floral honey versus honeydew honey, and season of production.
Botanical origin Botanical origin reveals that the leading Louisiana honey plants are TriJbliurn repens, Rubus, Berchernia scandens, and Salix. They are the major nectar contributors to the honey. Trifolium repens, Rubus, Berchemla scandens, and Salix also have the highest frequency of appearance in Louisiana honey samples. Miscellaneous Compositae (Asteraceae), Sapium sebiferum, Glycine max, Parthenocissus quinquefolia, and Cephalanthus occidentalis are confirmed as important nectar contributors in isolated samples. Miscellaneous Rosaceae (except Rubus) and Rhus radicans are notable for their frequency of appearance. Four plants worthy of recognition did not appear in Oertel's (1955) most recent list of Louisiana honey plants: Sapium sebiferum, Glycine max, Parthenocissus quinquefolia, and Rhus radicans. Sapium sebiferum is an introduced tree. The results indicate that Sapium sebiferurn is now widespread, especially in south-western Louisiana. Increased soybean production could easily account for Glycine max pollen appearing in the pollen spectrum. The Louisiana State Department of Agriculture (1966) stated that the soybean industry has made much progress in the last ten years;
State 0.02 0.03 12.9 0.01 1.8 0.09 0.01 0.2 0.2 0.01 O.2 0.5 0.4 0.02 0.2 0.3 !.0
A MELISSOPALYNOLOGICALSTUDY OF 54 LOUISIANA(U.S.A.) HONEYS
121
in 1965 there were 622,000 acres (more than any other crop) of soybeans grown in Louisiana. It is probable that Rhus radicans was not reported in 1955 because commercial beekeepers hesitated to admit that poison ivy is significant among the nectar-secreting flora. Other kinds of plant pollen appearing in the samples but not in Oertel's (1939) list of honey and pollen plants of Louisiana are: Allium, Chenopodium, Cocculus carolinus, Gramineae (Poaceae), Liriodendron tulipifera, Magnolia, Mimosa strigillosa, Myrica, Pinus, Rumex, Sagittaria, Sambucus, Scrophulariaceae, Umbelliferae (Apiaceae). All except Mimosa strigillosa generally appeared as trace pollen (less than 1%). It is assumed that the pollen grains from wind-pollinated plants such as Gramineae (Poaceae), Rumex, Pinus, Quercus, and Sagittaria are accidental honey contaminants. The pollen grains may have fallen or been blown into the nectar or honey; or were deliberately collected by the bees, stored in pollen cells, and entered the honey during the extraction process. Louisiana plants reported as honey or pollen plants by Oertel (1939), but whose pollen did not appear in the honey samples are: Ailanthus altissima, Aralia spinosa,
Bumelia, Citrus, Cliftonia, Convolvulus, Corculum leptopus, Cyrilla, Gossypium, Halesia, Ipomoea, Jacquemontia, Jussiaea, Melia azedarach, Oxydendrum arboreum, Perilla frutescens, Phoradendron flavescens, Richardia scabra, Sabal minor, S tellaria media, S tizolobium deeringianum, Tecoma radicans, Tilia, Vaccinium, Vigna sinensis, and Zanthoxylum clava-herculis. Oertel (1967, p. 10) helps account for the absence of Gossypium pollen in his statement, "Plant breeders have introduced a nectarless cotton so that destructive insects will not be attracted to the plant". Honey pollen analysis results concerning botanical origin did not consistently agree with beekeepers' reports. For example, sample 71 was reported as palmetto (Sabal minor) honey; this sample contained no Sabal minor pollen but had Rubus as dominant pollen and Trifolium repens as a secondary pollen. Louisiana honeys had fewer pollen grains per 10 g sample (Fig.l) than foreignproduced honeys. Over 50% of the Louisiana honeys had less than 20,000 grains per 10 g sample, whereas European workers (Maurizio, 1951) generally report that most honeys cleanly extracted by rotary extractor contain 20,000-100,000 grains per 10 g sample. European workers report that pollen content is increased in pressed or run honeys and is decreased when honey is adulterated with artificial products or by feeding sugar syrup to bees. In addition, the morphological structure of plants producing honey could affect the total pollen grain content. Louisiana beekeepers possibly produce less pollen-contaminated honeys than do European beekeepers, by using more refined techniques in their beekeeping and more efficient and cleaner extracting methods. Louisiana beekeepers may feed their bees more frequently, and thereby produce in some instances adulterated honeys with resulting lower pollen content. Oertel (1955) reported that nearly every beekeeper at some time or other feeds one or more colonies of bees.
122
MEREDITH HOAG LIEUX
The results presented in Table I indicate that the unifloral honeys of Trifolium repens, Berchemia scandens, and Rubus are to a considerable extent poor in pollen (< 20,000 pollen grains per 10 g). The Salix honey samples are for the most part richer in pollen (20,000-100,000 grains). No corrections were made to reestablish the probable nectar proportion associated with each pollen type as did Maurizio (1955, 1958) and Demianowicz (1964). Maurizio (1958) found most unifloral Trifolium repens samples in the 20,000-100,000 grain range. She .also found that three of five Salix samples had 20,000-100,000 grains; the other two Salix samples had over 100,000 pollen grains. Demianowicz (1964) reported that samples of unifloral Trifolium repens honey (samples obtained under controlled conditions) contained < 20,000 grains per 10 g sample. He also found that samples of Rubus honey were in the 20,000-100,000 grain range. A unifloral sample of Compositae honey (not shown in Table I) had the highest number of pollen grains per 10 g of any sample studied (approximately 3,300,000). This sample was probably polluted with pollen from pollen cells during the extracting process. Demianowicz (1964) reported that various Compositae (Asteraceae) honeys had total pollen grain counts per 10 g samples ranging from 1,950-43,480. A unifloral sample ofSapium sebiferum honey (not shown in Table I) had approximately 2,850 pollen grains per 10 g sample. The relatively large size of the Sapium sebiferum grain (33-36 # in diameter) possibly explains the low pollen count. Demianowicz (1964) concludes that there is an apparent relationship between the richness in pollen and the size of the grains: as a general rule, the smaller the pollen, the richer the honey. Louisiana honeys do not appear to have a particularly diverse botanical origin. Approximately 37% of the 54 Louisiana honey samples originated from six to ten different plant types. When combined, 75.9% of the samples originated from six to fifteen different plant types. Most of the Louisiana honey samples studied fell into the lower limits of Louveaux and Vergeron's 1964 study as to the pollen type ranges of Spanish honeys. For example, only 10% of Louisiana samples were in the 21-25 pollen type range.
Geographical origin Honeys collected from different honey producing areas of Louisiana are not easily distinguished from one another geographically by pollen types. Area 1 occupies chiefly the Mississippi River floodplain and delta and the floodplain of the Red River. The pollen types from honey gathered here indicate a varied flora of native and cultivated plants. The pollen assemblage represents plants growing on alternating low and higher ground with varying drainage, along streams, in woods, in swampy areas, in pastureland, and in cultivated fields. More honey is produced in this region than in any other area of the state. Three pollen types characteristic of Area 1 are: Cephalanthus occidentalis which indicates a swampy region, Mimosa strigillosa, and Forestiera
A MELISSOPALYNOLOGICALSTUDY OF 54 LOUISIANA (U.S.A.) HONEYS
123
acuminata which according to Brown (1945) is widely distributed in the Mississippi floodplain and along small streams and swamps. Maurizio and Louveaux (1965) reported that Mimosa pollen if found in honey could be used to distinguish exotic imported honeys fro_m central European honeys. Only one honey sample was collected from the pine-oak-hickory uplands of Area 2. Its pollen spectrum indicated no outstanding differences from the pollen assemblage of Area 1. This honey sample may be from an apiary located in a fringe area. Repeated attempts to collect more samples from area 2 were unsuccessful. Beekeepers living in this area reported one of two things: that they kept their bees in some other area or that the insecticide sprays used on cotton killed the bees and thus made commercial beekeeping unprofitable. Area 3 consists of pine-oak-hickory uplands and longleaf pine flatwoods. All samples collected in this area contained a characteristic pollen combination of Castanea and flex. There were no characteristic pollen types noted in either the prairie of area 4 or the pine-oak-hickory uplands and longleaf pine flatwoods of area 5. Area 6, the coastal marsh, is a low, wet plain along the Gulf of Mexico. For the most part, it is not suitable for beekeeping. Unifloral versus mixed honeys Of the 54 samples 31 were unifloral honeys (nine Trifolium repens, nine Rubus, seven Berchemia scandens, four Salix, one Sapium sebiferum, and one Compositae). All of the area 5 samples were unifloral honeys. There are probably more unifloral honeys produced in Louisiana than mixed honeys.
Floral versus honeydew honeys Most of the samples examined were clear colored and from nectar of flowers rather than from honeydew. But sample 40 (reported as a summer or fall honey; not included in the 54 samples from which data was recorded) may have originated, at least in part, from honeydew. Numerous fungus spores were observed. Zander (1949), as well as others, has reported that fungus spores are characteristic of honeydew honeys. Castanea pollen appeared as the most abundant pollen in sample 40. Barbier (1958) reported that Castanea produced both floral and honeydew honey that is rich in pollen. But Louisiana summer honeys often take up water and become dilute enough to ferment (K. Tucker, personal communication, 1971). Such conditions could account for sample 40 containing yeasts and fungus spores without the sample having necessarily derived from honeydew.
124
MEREDITH HOAG LIEUX
Season o f production Most samples examined were spring-produced honeys as indicated by their spring pollen spectra; a few were mixed spring and summer (e.g. samples 14, 26, 27, 38, 41, 76, 106, and 107); and possibly one sample was mainly summer or fall honey (sample 112). The pollen types used to identify late spring, summer, or fall honeys include: Glycine max, Cassia fasciculata, Polygonum, Lagerstroemia indica, L ythrum, Ampelopsis, Trachelospermum difforme, and others. The commercial beekeepers o f Louisiana probably sell mainly spring honey.
ACKNOWLEDGEMENTS I wish to recognize Dr. Clair A. Brown for his work on the photographs used in this manuscript and for the reference pollen he made available to me. I want to thank Dr. Shirley Tucker and Dr. Abdel Wahab. I. Allam (A. I. Allam) for reading the manuscript and giving technical advice. Due acknowledgement is made to National Science Foundation (U.S.A.) for their financial support during this study.
REFERENCES Barbier, E., 1958. Examen pollinique de quelques miels unifloraux. Ann. Abeille, 1: 73-76. Brown, C. A., 1945. Louisiana Trees and Shrubs. Louisiana For. Comm. Bull., 1:262 pp. Demianowicz, Z., 1964. Characteristik der Einartenhonige. Ann. Abeille, 7: 273-288. Erdtman, G., 1952. Pollen Morphology and Plant Taxonomy. (An Introduction to Palynology, 1 Angiosperms). Almquist and Wiksell, Stockholm, 539 pp. Louveaux, J. and Vergeron, Ph., 1964. Etude du spectre pollinique de quelques miels espangnols. Ann. Abeille, 7: 329-347. Maurizio, A., 1951. Pollen analysis of honey. Bee World, 32: 1-5. Maurizio, A., 1955. Beitr~igezur quantitativen PoUenanalyse des Honigs. Z. Bienenforsch, 3: 32-39. Maurizio, A., 1958. Beitr~igezur quantitativen PoUenanalyse des Honigs. Ann. Abeille, 1: 93-106. Maurizio, A. and Louveaux, J. 1965. Pollens de plantes Melif~res d'Europe. Union des Groupements Apicoles Fran~ais, Pads, 148 pp. Oertel, E., 1939. Honey and Pollen Plants of the United States. U.S. Dept. Agricult. Circ., 544, 63 pp. Oertel, E., 1955. The Beginner Beekeeper in Louisiana. Louisiana State Dept. Agricult. Immigr. Baton Rouge, La., 3rd ed., 46 pp. Oertel, E., 1967. Nectar and pollen plants. U.S. Dept. Agricult. Handbook, 335: 10-16. The Louisiana State Department of Agriculture, 1966. Louisiana-A Few Concrete Facts. Louisiana State Dept. Agriculture, Baton Rouge, La., 75 pp. Zander, E., 1935. Beitr~igezur Herkunftsbestimmung bei Honig. Pollengestaltung und Herkunftsbestimmung bei Bliitenhonig. Reichsfachgruppe linker, Berlin, 1:343 pp. Zander, E., 1949. Beitrage zur Herkunftsbestimmung bei Honig. Studien zur Herkun]tsbestimmung bei Waldhonigen. Franz Ehrenwirth, Miinchen, 4:267 pp.