Gretchen D. Jones

The use of ETOH for the dilution of honey

Since the beginning of honey production, certain honey types are preferred because they taste better, are better for cooking, or do not rapidly crystallize. Pollen found in honey is used to determine the honeys type. Techniques used to extract pollen from honey vary in the amount of honey examined, the amount of water used to dilute the honey, and the time and speed of centrifugation. These variations address the disparity in pollen recovery that is due to the specific gravity and sinking rates of the different pollen types. Ethyl alcohol (ETOH) was used as a dilution liquid for honey and compared to two water-dilution techniques, one with a short centrifugation time (1 minute) and the other using a long centrifugation time (10 minutes). The ETOH samples were centrifuged for 3 minutes. All samples were centrifuged at 4000 r.p.m. Significantly higher pollen concentration values were found in the ETOH-diluted samples. Pollen concentration values of the ETOH-diluted samples were 5.26 times greater than water-diluted, short centrifugation samples, and 3.26 times greater than water-diluted, long centrifugation samples. ETOH-diluted samples produced 16% more taxa than the water-diluted short centrifugation technique, and 10% more taxa than the water-diluted long centrifugation technique. The pollen spectra were not consistent across the three techniques. Additional research is needed to determine the efficiency of other ETOH concentrations in recovering pollen from honey. We recommend that the ETOH-dilution technique become the standard technique for the extraction of pollen from honey for pollen analyses because of the improved pollen recovery.

A comparison of pollen counts: Light versus scanning electron microscopy

Palynologists use compound light microscopy (LM) for pollen identification and interpretation and scanning electron microscopy (SEM) for morphological comparisons and taxonomy. As we are unaware of any published reports comparing LM and SEM pollen counts and identifications of the same sample, we decided to examine a surface soil sample using both microscopes. Standard palynological extraction techniques were used. Two, 300 grain counts were made using LM, and two, 300 counts with SEM. Pollen grains viewed with SEM were also divided into three categories, “identifiable,” “obscured,” and “virtually impossible to identify”. Eighty‐six (86) percent of the pollen grains counted with SEM were classified as “identifiable” or “obscured.” Pollen concentration values ranged from 385,714 (LM Count #2) to 900,000 (SEM Count #1) per gram of soil. Regardless of the microscope used, Ligustrum spp., Myrtaceae‐type, and Tilia spp. had the greatest number of pollen grains. A total of 73 taxa were found. A scan of the unexamined portion of the stubs resulted in 20 additional taxa. There were no significant differences between the counts made with the two microscopes (ANOVA, p>0.05, F = 0.18, df = 76). However, there were significantly more taxa found with SEM than with LM (t‐test, T = 0.05). Sample preparation and the time needed to count, analyze, photograph and print the micrographs are the same regardless of the microscope used. The sample, information needed, and funding will determine which technique to use.

COTTON POLLEN RETENTION IN BOLL WEEVILS: A LABORATORY EXPERIMENT

Abstract Cotton pollen is thought to exist in the gut of boll weevils for at least 24 hours. Because finding whole or broken cotton pollen grains are valuable clues in determining when boll weevils were living in cotton, the retention of cotton pollen in the gut of the weevil was examined. Boll weevils were fed cotton buds, cotton flowers, and water for five days, then they were placed into cleaned cages without any food. However they were allowed to drink water freely. Weevils were examined at 0, 24, 48, 72, 96, and 120 hours after the buds and flowers were removed. One hundred weevils were dissected at each time interval, and each gut was placed onto a glass slide. Light microscopy was used to determine the presence or absence of cotton pollen. More weevils (63%) at 0 hours contained pollen than weevils at any other interval. Whole pollen grains were not found in weevils after 24 hours. The 120 hour interval contained a single weevil with pollen. In the laboratory test, whole cotton pollen grains indicated that feeding on cotton occurred within 24 hours. When only halves and fragments of pollen were found, cotton feeding occurred 48–120 hours earlier. Additionally, when six or more cotton pollen grains were found in a weevil, it had fed on cotton within 24 hours. This information narrows the boll weevil dispersal times and distances from cotton fields and overwintering areas.

ALMOND, MELON, AND PIGWEED POLLEN RETENTION IN THE BOLL WEEVIL (COLEOPTERA; CURCULIONIDAE)

Abstract The boll weevil (Anthonomus grandis Boheman) remains a devastating insect pest on cotton (Gossypium hirsutum C. Linnaeus), particularly where it has not been eradicated. Identifying and understanding the survival of overwintered boll weevils, when cotton is not available, is important in designing mitigation programs and controlling this insect pest, especially in areas where boll weevils are active all year. Many non-malvaceous taxa have been listed as possible overwintering adult food sources. The purpose of this research was to determine if pollen from three non-malvaceous taxa could be used as food sources for overwintering boll weevils in the Lower Rio Grande Valley of Texas, where weevils are active year-round and during cotton-free periods. The taxa are almond [Prunus dulcis (Mill.) D.A. Webb], melon (Cucumis melo C. Linnaeus subsp. melo), and pigweed (Amaranthus sp.), Furthermore, it was intended to determine the retention of these pollen types in the boll weevil gut. More weevils (84%) contained almond pollen when they had additional water than those (30%) that did not have additional water. Almond pollen was found in 100% of the examined weevils after 72 hours, and in 80% after 96 hours. Pigweed and melon pollen were never found after 48 hours. Overall, almond pollen was found in more weevils (96%) than pigweed (40%), or melon (12%). In addition, a greater number of almond pollen grains (539) were found than pigweed (41), or melon (21). This shows that these types of pollen were consumed, and could play a part in the survival of overwintering weevils. However, more research is needed to examine the nutritional value of these, and other, pollen grains for overwintering boll weevil survival, and to determine the length of time weevils can survive feeding on these pollen types.

Pollen Studies of East Texas Honey

Since the beginning of honey production, certain honey types have been favored because they taste better, are better for cooking or do not rapidly crystallize. Thus, they are preferred over others, are in high demand and are sold at higher prices. The pollen of 37 honey samples from East Texas was examined. Pollen was recovered from the honey by using an alcohol-dilution method. Overall, 431 taxa identified into 61 families, 104 genera and 85 species were found in the samples. The number of taxa per sample varied from 17–52. Half of the samples contained 31–40 taxa, indicating a high diversity in botanical origin. Three taxa were found in >50% of the samples and are the most important: Berchemia scandens, Salix nigra and Toxicodendron radicans. Berchemia scandens was found in 89% of the samples and was a predominant type in three samples and an important secondary type in 14. Both Salix nigra and Toxicodendron radicans pollen occurred in 83% of the samples and neither occurred as a predominant or secondary type. Three samples were Berchemia scandens unifloral honey. By examining the pollen in honey, it can determined which habitats honeybees visit, which plants honeybees use as food, if they visit row crops and orchards and their role in pollination. In order to differentiate honey from the United States of America (USA) from honey produced in other countries, the honey from each state must be analyzed. Only by analyzing the pollen in the honey of the USA can it be reliably differentiated from foreign honey that is being sold as produced in the USA.

Pollen analyses of tarnished plant bugs

Wild host plants are important for tarnished plant bug (TPB — Lygus lineolaris) populations, especially when cultivated crops are not flowering. Knowledge of native habitats is important for managing this insect pest. Although pollen has been used to characterize dispersal and food sources of many insects, it has not been used for TPB. The purpose of this study was to determine if pollen analyses could be used as a tool to determine the non-crop plants associated with TPB. Thirty-eight TPB were collected from light traps set up at two sites near the USDA-ARS research farm at Stoneville, Mississippi on 27 June 2011. Overall, 1183 pollen grains from 79 plant taxa were found in the samples. The pollen recovered was identified to 29 families, 33 genera, and 24 species. Many of the taxa of pollen found, such as Saururus cernuus C. Linnaeus (lizards tail), Sagittaria, Echinodorus and Carya aquatic (water hickory), occur in wet habitats. From the assemblage of identified pollen grains, TBP from both sites utilized plants from disturbed and/or wet habitats.

Forensic pollen geolocation techniques used to identify the origin of boll weevil re-infestation

Abstract The boll weevil, Anthonomus grandis, entered the United States of America in the early twentieth century and became a major pest in cotton, Gossypium spp. Shortly after the passage of Tropical Storm Erin on 16 August 2007 through the South Texas/Winter Garden boll weevil eradication zone, over 150 boll weevils were captured in the Southern Rolling Plains (SRP) eradication zone that was essentially weevil-free since 2003. Pollen analyses were made of the SRP weevils and weevils collected in two suspected source zones, Cameron (Southern Blacklands eradication zone) and Uvalde (Winter Garden eradication zone). An additional examination of the palynological evidence and examination of additional pollen residue shed new light on this event and strengthens the conclusion that the Uvalde area was the source of the SRP weevils. A total of 192 pollen grains from 39 taxa were found in the SRP weevils: 1904 pollen grains from ten taxa from the Cameron weevils and 148 grains from 28 taxa in the Uvalde weevils. The SRP weevils shared 16 taxa, including Phermeranthus sp. (flameflower) with the Uvalde weevils and only five taxa with the Cameron weevils. Common taxa between SRP and Uvalde weevils and the lack of the dominant ‘low spine’ Asteraceae that occurred in all Cameron samples confirm that the SRP weevils originated from the South Texas/Winter Garden zone. Problems associated with this type of research are similar to those in forensic palynology. These problems include the unknown origin of the weevils, pollen contamination and care and storage of the samples.

Pollen analysis of Agathirsia wasps

Abstract Parasitic wasps are one of the most abundant natural enemies of insect pests and the most ecologically understudied. Agathirsia is a genus of solitary, parasitic wasps, restricted to arid regions of the south-western United States of America and Mexico. They often parasitise the larvae of lepidopteran (butterflies and moths) crop pests. Pollen analyses were conducted on 127 specimens of 19 Agathirsia species to determine possible food sources and plant taxa important to these beneficial insects. Pollen was examined with both light microscopy (LM) and scanning electron microscopy (SEM). Pollen was removed from the head more than any other insect part (96% of the time). In the LM analyses, nearly 1700 pollen grains were recorded and assigned to 117 pollen types, 49% of which were Asteraceae. For the SEM analyses, pollen was identified but not counted. Eighty-eight pollen types were identified, of which 48% were Asteraceae. Pollen from 36 families, 65 genera and nine species were identified. Asteraceae pollen was classified into more genera than any other plant family. Common taxa included Ambrosia, Artemisia and Baccharis. The large diversity and high number of Asteraceae indicates that these plants can help sustain Agathirsia populations near crop fields, and ultimately reduce the amount of applied pesticides and save farmers time and money.

Pollen contamination of boll weevil traps

Abstract The boll weevil, Anthonomus grandis Boheman, has been the most devastating insect pest of cotton, Gossypium hirsutum L., in the southern United States of America (USA). Although thought to feed only on cotton, the list of non-cotton alternative food sources increases yearly. Many of these taxa are thought to be contaminates and not food sources. The purpose of this research was to examine the possibility that weevils become contaminated with pollen while sitting in the trap. Between January and April, boll weevil traps were placed near Brownsville and Weslaco (a substitute location) and in Santa Ana Wildlife Refuge, Texas. Pollen was removed from the traps pole, skirt, mesh cone, and lid by wiping them with an individual, sterile, 100% cellulose acetate filter. The original trap was replaced with a replacement trap that was left for three days, then wiped for pollen. Little pollen was found on the trap parts regardless of the month or the site. The mesh cone was the least contaminated. From these data, it is doubtful that weevils become contaminated with pollen while sitting in the trap. Additional research is needed to examine if pollen is transferred among trapped weevils.