Manú National Park is a UNESCO Biosphere Reserve nested within the most biodiverse region in the world: the tropical Andes (Myers et al. 2000). Manú Park, and its surrounding forests encompass a remarkable elevational gradient (hereafter the 'Manú Gradient') of over 3000 m, reaching from the lowland Amazon rainforest to the Puna grasslands of the high Andes. The Manú Gradient has a rich history of ornithological research (discussed in Walker et al. 2006), and over the last decade the Manú Gradient has been the focus of numerous ornithological studies as part of the Manú Bird Project (e.g. Merkord 2010, Jankowski et al. 2012a, Jankowski et al. 2012b, Londoño et al. 2014, Londoño et al. 2016, Dehling et al. 2014, Munoz 2016). Along the gradient, tree composition and forest structure have also been described (e.g. Jankowski et al. 2012b, Malhi et al. 2010, Hillyer and Silman 2010). The wide interest in the avian community of Manú make it an ideal system for studying hummingbird pollination: population structure, range limits, and locations of uncommon and understudied hummingbirds are described and published. For example, focused studies of the high elevation Shining Sunbeam (Aglaeactis cupripennis) have demonstrated the effectiveness of the Manú Gradient as a study system for bird pollination (Hazlehurst et al. 2016, Hazlehurst and Karubian 2016). Therefore, the objectives of this study were to, 1) document the occurrence of putatively bird pollinated plants with voucher specimens along the Manú Gradient, 2) describe the occurrence and diversity of hummingbirds using mist-net surveys, and 3) record hummingbird visitations to flowering plants.

Site Selection

We surveyed three field sites spanning an altitudinal gradient of 2400 m (400 m to 2800 m) in the southeastern Andes (Table 5): La Estación Biológica Wayqecha (Paucartambo Province, Cuzco Region, 2800 m), San Pedro (Paucartambo Province, Cuzco Region, 1400 m), and Pantiacolla (Manú Province, Madre de Dios Region, 400 m). This area is one of the most biologically rich regions in the world with an estimated species pool of nearly 1100 birds (Walker et al. 2006). To our knowledge, a comprehensive survey of the vascular plants of the region does not exist, although an increasing number of plant identification resources for this region are being made available by the Field Museum of Natural History (http://fieldguides.fieldmuseum.org). Wayqecha is characterized as high elevation cloud forest, with a mosaic of mature forest and areas with shorter trees and woody shrubs that transitions into puna grassland above treeline. San Pedro is predominately mid-montane humid rainforest, but also includes the lower extent of the montane cloud forest. Pantiacolla is situated at the interface between the Andean foothills and the lowland Amazon. Detailed environmental characteristics for these sites have been summarized in Malhi et al. (2016). Sampling was carried out between September 4, 2016 and October 13, 2016, falling within the avian montane breeding season. A distinct rainy season occurs from November through April and a dry season from May through August. Annual precipitation for higher elevations (2700-3000 m) ranges from 1700-2000 mm (Girardin et al. 2010) and is generally >2000 mm for lowland (100-400 m elevation) sites (Rapp and Silman 2012). Time constraints afforded less than two weeks (10.3 ± 2.1 days) for botanical and avian sampling at each site.

Data Collection

Pre-cut singletrack trails were used to access sampling areas away from the Manú Road (main access road that runs along the southeastern border of Manú National Park). We sampled hummingbirds using standard (12 x 3 m, 34 mm mesh) mist-nets along trail systems only. Mist-netting sites were sampled during the primary breeding season (August–November) for two consecutive days from approximately 0600–1200 hrs during suitable weather conditions (i.e., no periods of extended heavy rain, high winds, or other situations that could compromise researcher or bird safety). Each site consisted of an array of ten to fifteen nets placed in forested and open habitat and spaced at intervals of 25-50 m. Ten sites were sampled at Wayqecha and San Pedro, and 8 sites were sampled at Pantiacolla. Hummingbird bill length was measured from the bill tip to the nares. Bill width was measured from the anterior edge of the nares. All captured hummingbirds were marked by cutting the terminal 1-2 cm of one rectrix to avoid resampling of individuals.

Both trails and the Manú Road were used to opportunistically collect plants. Plants were considered putatively bird pollinated if they met criteria adhering to typical bird pollination ‘syndromes’; namely, dilute nectar and long tubular flowers (Fenster et al. 2004), though we acknowledge the limitations of surveying by these critera (Ollerton et al. 2009). Plants of interest were photographed, their location marked using a hand-held Garmin 64s global positioning system, and a description of the immediate habitat recorded. We then measured nectar concentration of mature flowers (Sper Scientific no. 66214-988), recorded corolla dimensions and colour (by visual inspection), and processed each plant using standard herbarium techniques Bridson and Forman 2000) (SERFOR collection permit no. 343-2016). All dried and pressed specimens are deposited at the herbaria of the Universidad Nacional Agraria La Molina (MOL), Peru and the University of British Columbia (UBC), Canada (SERFOR export permit no. 09125-2017).

We identified 44 putatively bird pollinated plants of interest belonging to 16 families (Table 1, Figs 1, 2, 3, 4, 5). Corolla length and width of sampled plants ranged from 8-120 mm (x̅ = 39.7 ± 27.4, n = 42) and 1-60 mm (x̅ = 11.4 ± 12.2), respectively. We measured nectar concentration for 11 of these species. In each case, nectar concentrations fell within a typical bird pollination syndrome (Stiles 1978, Fenster et al. 2004), ranging from 12-25.5% (Table 1). Corolla colour and immediate habitat characteristics were recorded for each plant (Table 1, see also Table 5).

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b

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d

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Figure 1.

Diversity of putatively bird pollinated plants of the Manú Gradient. MMAB collection numbers listed in Table 1.

a: MMAB 1 (Besleria sp. 1)  
b: MMAB 2, 3 (Centropogon granulosus)  
c: MMAB 6, 7 (Miconia sp. 1)  
d: MMAB 8, 9 (Columnea aff. shimpfii)  
e: MMAB 20, 21 (Sanchezia sp. 1)  
f: MMAB 22, 23 (Columnea guttata)  

a

b

c

d

e

f

Figure 2.

Diversity of putatively bird pollinated plants of the Manú Gradient. MMAB collection numbers listed in Table 1.

a: MMAB 24, 25 (Heliconia subulata)  
b: MMAB 26, 27 (Guzmania weberbaueri)  
c: MMAB 39, 40 (Drymonia urceolata)  
d: MMAB 45 (Centropogon congestus)  
e: MMAB 48, 49 (Oreocallis grandiflora)  
f: MMAB 50, 51 (Siphocampylus scandens)  

a

b

c

d

e

f

Figure 3.

Diversity of putatively bird pollinated plants of the Manú Gradient. MMAB collection numbers listed in Table 1.

a: MMAB 52, 53 (Siphocampylus orbignianus)  
b: MMAB 54, 55 (Brachyotum rostratum)  
c: MMAB 56, 57 (Aetanthus nodosus)  
d: MMAB 58, 59 (Gaultheria sp. 1)  
e: MMAB 60, 61 (Passiflora mixta)  
f: MMAB 62, 63 (Fuchsia sp. 1)  

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b

c

d

e

f

Figure 4.

Diversity of putatively bird pollinated plants of the Manú Gradient. MMAB collection numbers listed in Table 1.

a: MMAB 64, 65 (Desmodium sp. 1)  
b: MMAB 66, 67 (Siphonandra sp. 2)  
c: MMAB 68, 69 (Bomarea sp. 1)  
d: MMAB 70, 71 (Drymonia semicordata)  
e: MMAB 74, 75 (Costus sp. 3)  
f: MMAB 76, 77 (Heliconia densiflora)  

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b

c

d

e

f

Figure 5.

Diversity of putatively bird pollinated plants of the Manú Gradient. MMAB collection numbers listed in Table 1.

a: MMAB 78, 79 (Costus sp. 2)  
b: MMAB 80, 81 (Besleria sp.4)  
c: MMAB 82, 83 (Heliconia schumanniana)  
d: MMAB 84, 85 (Heliconia lingulata)  
e: MMAB 86, 87 (Besleria sp. 2)  
f: MMAB 92, 93 (Pentagonia sp. 1)  

We recorded 23 hummingbird visitations to 18 plant taxa belonging to 12 plant families (Table 2). Bill length and width of sampled hummingbirds ranged from 11.5-39.6 mm (x̅ = 24.3 ± 7.6, n = 41) and 2.5-3.0 mm (x̅ = 2.9 ± 0.4, n = 40), respectively (Figs 6, 7, 8, Table 3).

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b

Figure 6.

Representative hummingbirds captured near San Pedro (Paucartambo Province, Cuzco Region, 1400 m). Photo "b" taken by Meredith Miles, Wake Forest University.

a: Booted Racket-tail (Ocreatus underwoodii)  
b: Speckled Hummingbird (Adelomyia melanogenys)   

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b

Figure 7.

Representative hummingbirds captured near La Estación Biológica Wayqecha (Paucartambo Province, Cuzco Region, 2800 m).

a: Shining Sunbeam (Aglaeactis cupripennis)  
b: Collared Inca (Coeligena torquata)  

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b

Figure 8.

Representative hummingbirds captured near Pantiacolla (Manú Province, Madre de Dios Region, 400 m).

a: Blue-fronted Lancebill (Doryfera johannae)  
b: White-necked Jacobin (Florisuga mellivora)  

Diversity of plants exhibiting the bird pollination syndrome does not differ across the gradient in the time frame sampled (Table 4).

Hummingbird pollination is common and well-established in Neotropical montane and lowland environments. Our observations and collected specimens exemplify that bird-plant interactions are readily observed along the Manú Gradient - an area that is relatively accessible has been subject to only a handful of studies on hummingbird pollination (Oreocallis grandiflora, Proteaceae; Hazlehurst et al. 2016, Hazlehurst and Karubian 2016).

Along the gradient, putatively bird pollinated plants were generally characterized by long corollas and were predominantly coloured red, yellow, orange, or some combination thereof. Previous documentation of bird pollination exists for each of the 16 families collected (Cronk and Ojeda 2008, Johnson and Nicolson 2008), but undocumented species-level bird pollination systems may arise from focusing on lesser-stuided taxa (e.g. Thyrsacanthus, Pentagonia, Pachystachys). Many putatively bird pollinated plants contained too little nectar to effectively measure sugar concentration at the time of sampling. We suspect that early morning visitations by nectarivorous birds and insects (i.e., both pollinators and nectar robbers) influenced this outcome. Indeed, in some cases inspection of certain plants revealed that the flower had been recently robbed as indicated by punctures at the base of the corolla. In as little as bird pollination has been studied along the Manú Gradient, even less is known of the ecological and evolutionary dynamics of nectary robbery. As this survey was preliminary, time did not allow for multi-day sampling at one locale to isolate nectar. A focus on a specific plant taxon would allow familiarity for nectar phenology and hence, more effective collection of nectar.

We recorded 23 independent visits by hummingbirds to 19 different plant taxa over 33 days. These observations by no means represent a comprehensive list of the total diversity for hummingbirds (Walker et al. 2006), bird-pollinated plants, or the interactions between these two groups. An estimate of total diversity will come only with an extended sampling effort at each site. Relatively few hummingbirds were captured or observed in the lowlands (Table 3) compared to the other two sites. It is likely that this resulted from differences in foraging behavior between hummingbird species, rather than local abundance. For example, in the lowlands, a higher proportion of hummingbirds (e.g. Phaethornis) exhibit traplining behaviour (i.e. repeated visits along a route of flowering locations) compared to territorial guarding of floral resources. In addition, because of the higher canopy, many of the trees, lianas, and epiphytes inhabit canopy heights that are logistically difficult to sample.

The number of plants exhibiting bird pollination syndrome and number of bird visits observed are comparable between sites. That is, at a coarse scale we did not find any indication that elevation affects the absolute diversity of bird pollinated plant taxa (as expected by Cruden 1972), although the Manú Gradient would be an ideal location to test the hypothesis that bird and insect pollinated plants occupy distinct ecological niches. Between species, corolla length and width encompasses a great amount of variation, but hummingbird bill morphology varies less Tables 1, 3. This may speak to the adaptability of flowers relative to bills. It may be that because flowers serve a singular purpose (attraction and exclusion of pollinators and robbers, respectively), whereas bills have many uses (feeding, aggression, preening, balance), that bill evolution is relatively constrained. Bill morphology data will be used to inform phylogenetic tests of bill-flower shape evolution in future studies.

Evaluating the extent to which plants and their pollinators contribute to maintaining local biodiversity, and identifying keystone species within these systems (Ebenman and Jonsson 2005) will be important to maintaining ecological and cultural heritage in the Manú region (Ministerio del Ambiente (Ministry of Environment) 2017). This study provides a baseline for future work in pollination ecology along the Manú Gradient. Any one of the 44 plant species highlighted here warrants closer investigation, and we anticipate that further studies will help clarify the roles of hummingbirds as pollinators for the plant taxa described herein..

The authors would like to acknowledge Daniela Olivera (Universidad Nacional de San Antonio Abad del Cusco) for their assistance in the field, as well as the staff of Wayqecha Biological Station, Cock-of-the-Rock Lodge, and Pantiacolla Lodge. We also acknowledge Jenny Muñoz (University of British Columbia), the Asociación para la Conservación de la Cuenca Amazónica (ACCA), and the Servicio Nacional Forestal y de Fauna Silvestre (SERFOR) for assistance in administering research permits, and Prof. Carlos Reynel and the herbarium staff at Universidad Nacional Agraria La Molina for generously providing assistance at all stages of this project. Identification of our Geseneriacae specimens were greatly improved by Dr. John Clark (University of Alabama). Identification of numerous other specimens were assisted by Dr. Quentin Cronk (University of British Columbia). This expedition was supported by grants from the Natural Sciences and Engineering Research Council of Canada (NSERC) Discovery Grants Program, held by Dr. Quentin Cronk (RGPIN-2014-05820) and Dr. Jill Jankowski (RGPIN-2012-418294), as well as a Walter H. Lewis Award in Plant Biodiversity to MMA Boehm and a Werner and Hildegard Hesse Research Award to MN Scholer.