International Journal of Ecology

International Journal of Ecology / 2008 / Article

Research Letter | Open Access

Volume 2008 |Article ID 260754 |

Matthew A. Neatrour, Robert H. Jones, Stephen W. Golladay, "Assessment of Nutrient Limitation in Floodplain Forests with Two Different Techniques", International Journal of Ecology, vol. 2008, Article ID 260754, 4 pages, 2008.

Assessment of Nutrient Limitation in Floodplain Forests with Two Different Techniques

Academic Editor: Shibu Jose
Received29 Nov 2007
Accepted07 Apr 2008
Published15 May 2008


We assessed nitrogen and phosphorus limitation in a floodplain forest in southern Georgia in USA using two commonly used methods: nitrogen to phosphorus (N:P) ratios in litterfall and fertilized ingrowth cores. We measured nitrogen (N) and phosphorus (P) concentrations in litterfall to determine N:P mass ratios. We also installed ingrowth cores within each site containing native soil amended with nitrogen (N), phosphorus (P), or nitrogen and phosphorus (N + P) fertilizers or without added fertilizer (C). Litter N:P ratios ranged from 16 to 22, suggesting P limitation. However, fertilized ingrowth cores indicated N limitation because fine-root length density was greater in cores fertilized with N or N + P than in those fertilized with P or without added fertilizer. We feel that these two methods of assessing nutrient limitation should be corroborated with fertilization trials prior to use on a wider basis.

1. Introduction

Many ecologists have applied Liebig’s law of the minimum to forest ecosystems and have sought to determine what nutrient limits net primary production (NPP). Globally, NPP generally is considered to be limited by nitrogen (N) [1], but many forests may be switching from nitrogen to phosphorus limitation because humans have increased atmospheric nitrogen inputs to forests through fossil fuel combustion, nitrogen fertilizer production, and other human activities [2]. Assessing nutrient limitation, however, has been historically problematic because it typically has required laborious and time-consuming fertilization trials [3]. Within the last decade, ecologists have increasingly used simpler methods to determine nutrient limitation in forests, such as foliar N : P ratios [411] or root growth into fertilized ingrowth cores [1218], but no study to date has explicitly compared these two techniques.

The root ingrowth core method was originally developed to measure root production and involves excavating a soil core that is replaced with root-free growth medium (e.g., native soil, vermiculite, peat), sometimes enclosed in a mesh bag [18]. The core is extracted after a given period of time and roots growing into the core are removed. The ingrowth core technique was modified by Cuevas and Medina [13] to measure nutrient limitation by comparing root growth into cores fertilized with different nutrients. Their modification makes use of the well-known plastic response of plants to proliferate roots into nutrient-rich patches [19] and assumes that roots respond more strongly to limiting nutrients. Raich et al. [17] corroborated this technique in forests with known nutrient limitation, and it has recently come into widespread use as a result of their work [12, 1416, 18].

N : P ratios in plant tissue have been used as diagnostic indicators of N or P limitation, and thresholds have been established for some ecosystems [3, 9, 20]. The use of N : P ratios to determine N or P limitation is based on the assumptions that either N or P is limiting and that N or P concentration in plant tissue reflects nutrient deficiencies in soils. For wetland plant communities in Europe, Koerselman and Meuleman [3] suggested that communities with an N : P ratios > 16 were P-limited while those with an N : P ratios < 14 were N-limited. Lockaby and Conner [20] extended this work to forested wetlands of the southeastern United States by examining N : P ratios in litterfall and proposed that N : P ratios < 12 were N-limited and those > 15 were P-limited.

Here, we used the fertilized ingrowth core and litter N : P ratio techniques to assess nutrient limitation in a floodplain forest along a blackwater stream in southwestern, Ga, USA. Blackwater streams typically have lower inorganic materials and suspended sediments in their waters and therefore floodplain forests associated with these systems generally are thought to be nutrient poor [21]. In other blackwater systems in the southeastern United States, P is generally considered to be limiting [8, 22]. Thus, we expected both techniques would indicate P limitation.

2. Methods

Three 2 6 × 2 6  m plots were established within the floodplain forest of the Chickasawhatchee Wildlife Management area of Baker and Calhoun counties, Ga, USA. The overstory was dominated at all sites by Liquidambar styraciflua L, Acer rubrum L, and various Quercus species (Quercus lyrata Walt., Quercus laurifolia Michx., Quercus nigra L). The understory consisted primarily of Smilax spp., Toxicodendron radicans (L.) Kuntze and Sabal minor (Jacq.) Pers. Soils were Typic Alabaqualfs (Megget series) or Typic Fluvaquents (Muckalee series) [23, 24]. Mean annual temperature during the study year was 1 9 C and annual precipitation was 141 cm, which was 11% above normal (National Climatic Data Center, Asheville, North Carolina).

Forty cores of mineral soil (2-cm diameter × 20-cm deep) were collected with a soil probe from each plot. Soil was sieved through a 2-mm mesh screen to remove coarse fragments, combined by plot, and air dried. Soil pH was measured using an Accumet pH meter, and total C and N was determined using a Perkin Elmer Series II CHNS/O analyzer (Perkin Elmer Inc., Boston, Mass, USA). Soils were analyzed for extractable phosphate on a Lachat Quickchem AE autoanalyzer following double-acid extraction [25].

Five 0.25-m2 litter traps were placed randomly in each plot in September 2003, and litterfall was collected in November 2003 to determine litter N and P concentrations. Litter was dried to a constant mass ( 6 0 C), weighed, ground using a Wiley mill (Thomas Scientific, Swedesboro, NJ, USA), and pulverized with a ball Spex 8000D ball grinder (SPEX CertiPrep Group, Metuchen, NJ, USA). Litter N concentration was determined using the dry combustion method on a Perkin Elmer Series II CHNS/O analyzer. Litter P was measured with the dry ash method [26] followed by analysis on a Lachat Quickchem AE autoanalyzer (Lachat Instruments, Milwaukee, Wis, USA). Litter N : P ratios were calculated as mass ratios typically used for forest and wetland vegetation.

For the ingrowth core study, each plot was divided into 2 × 2  m grid cells consisting of 196 grid intersections and 44 of the intersections were designated randomly for installation of ingrowth cores. In early July 2003, a 10.16-cm diameter × 20-cm deep core was extracted at each selected location. The core was filled with soil collected from a nearby site that was sieved through a 2-mm screen to remove roots. Soil was packed to approximately the same bulk density as the soil that was removed. The core was either left unfertilized or had phosphorus, nitrogen, or phosphorus and nitrogen fertilizers mixed into the top 5 cm. Treatments were replicated 11 times per plot. Nitrogen was added as 0.75 g of POLYON coated urea (43-0-0 NPK, Harrell’s Inc., Lakeland, Fla, USA), and phosphorus was added as 0.40 g triple superphosphate (0-46-0 NPK, Southern States Cooperative, Christiansburg, Va, USA) to raise soil nutrient availability by 400 kg N/ha and 100 kg P/ha, respectively. Cores were harvested after 4 months in late November 2003 by extracting a 7.62-cm diameter × 20-cm deep core from the middle of each ingrowth core. Seven cores were not collected due to damage by animals. Cores were washed over a 1-mm mesh screen to remove adhering soil particles. Roots were separated from soil organic matter in the lab and subsampled for root length analyses. Each subsample was scanned and analyzed using WinRHIZO software (Regent Instruments, QC, Canada) to determine specific root length (SRL, cm of root per g of root). Root samples were dried to a constant mass at 6 0 C and weighed. Root length density (km root per m3 of soil) was calculated by multiplying SRL by root mass. We ran a one-way ANOVA to compare root length density among fertilization treatments with plots as blocking factors using the GLM procedure of SAS version 8 (Statistical Analysis System, Cary, NC, USA). We also used to 95% confidence limits to determine whether N : P were different from threshold ratios for 12 and 15, for N and P limitation, respectively.

3. Results and Discussion

The fertilized ingrowth core method indicated N limitation because root length density in cores fertilized with N or N + P was nearly double that of root length density in P-fertilized cores or in cores with no added fertilizer ( 𝑃 < . 0 1 , Figure 1). In addition, root length density did not differ between P-fertilized cores and cores without fertilizer ( 𝑃 < . 9 9 ) . According to 95% confidence intervals, litter N : P ratios (mean = 19.1) were greater than 12 but not than 15, the thresholds for N and P limitation, respectively, in floodplain forests of the southeastern United States [20]. This suggested P limitation or colimitation by N and P in this forest. Low N (0.8%) and P (0.04%) compared to other floodplain forests [20] may support colimitation (Table 1). However, high nutrient resorption proficiency of P, defined by Killingbeck [27] as the degree to which plants can reduce the level of a given nutrient in senescing leaves, lends support for P limitation in this forest. Proficiency was < 0.05% for P and > 0.7% for N, which suggested nearly complete resorption of P and intermediate resorption of N [27]. The discrepancy between the two techniques is somewhat puzzling. In the following paragraphs, we provide possible reasons for these contradictory results.


Extractable PO4-P ( 𝜇 g/g)1.0 ( 0 . 7 - 1 . 3 )
Total N (%)0.07 ( 0 . 0 6 - 0 . 0 8 )
Total C (%)1.6 ( 1 . 3 - 1 . 9 )
pH5.0 ( 4 . 2 - 5 . 4 )
P concentration (%)0.04 ( 0 . 0 3 - 0 . 0 5 )
N concentration (%)0.80 ( 0 . 6 7 - 0 . 9 8 )
C concentration (%)49.8 ( 4 9 . 2 - 5 0 . 8 )
N  :P ratios19.1 ( 1 6 . 1 - 2 1 . 7 )
C : N ratios63.7 ( 5 0 . 0 - 7 6 . 0 )

We feel there are three likely explanations why roots responded only to N fertilization in potentially P-limited conditions. First, the use of fertilized ingrowth cores to measure nutrient limitation is centered on the assumption that roots respond more strongly to limiting nutrients than to those that are “non” limiting. However, this assumption may not always be valid for all forests. Hodge et al. [28] suggested that proliferation may be an adaptive response by plants to obtain nutrients when in competition with other plants rather than a response based on immediate nutritional needs. If this is the case, plants may not respond more strongly to limiting nutrients because they are constrained by their evolutionary history. For example, plants species adapted to soils that are historically N-deficient only may have the capacity to proliferate roots into N-rich patches even when P is limiting. Second, the ingrowth core method operates on the implicit assumption that all plants within a community can proliferate roots into nutrient-rich patches. However, not all plants can proliferate roots [29], and nutritional status of individual species may differ from that of the whole plant community [30]. Therefore, proliferation may be a nutritional response by individual species that may or may not reflect nutrient limitation of the whole community. Third, P may have become rapidly unavailable to plants due to adsorption into Fe and Al complexes or leaching from the cores. The type of fertilizer used in our study, triplesuperphosphate (TSP), is water soluble and fast releasing even though the level of P we used was relatively high (100 kg/ha). Although this may help to explain the lack of response to P, it does not offer any insight as to why roots responded strongly to N fertilization.

There are also several reasons why litter N : P ratios may be poor indicators of P limitation in N-limited conditions. First, the use of N : P ratios to indicate nutrient limitation requires that either N or P limits NPP. N : P ratios cannot detect limitation by another resource (i.e., other nutrient, light, water) or colimitation by multiple resources. The fertilized ingrowth core as used in our study also could not determine nutrient limitation by other resources besides N and P but has the flexibility to at least test limitation or colimitation by other nutrients. Second, assessing nutrient limitation through N : P ratios also necessitates that these ratios are indicators of nutrient deficiencies in soils. However, N : P ratios of individual species do not always vary across a gradient of known nutrient limitation [31], but Aerts and Chapin [30] warned that N : P ratios of individual species may differ from community N : P ratios. Third, thresholds for N or P limitation often are hard to establish. Although there is a general consensus that low N : P ratios indicate N limitation, there is no agreement as to whether intermediate to high N : P ratios indicate P limitation [32]. Some researchers have found N limitation or colimitation by N and P when N : P ratios are high [9, 33]. Finally, all of the above problems may have been compounded by the use of litter N : P ratios instead of N : P ratios in live foliage because of differences in retranslocation efficiencies between N and P. The use of foliar N : P ratios in conjunction with litter N : P ratios may have provided more insight into what nutrient was limiting.

In conclusion, we found that these two techniques gave contradictory results as to what nutrient was limiting (i.e., N or P) and therefore cannot be used to reliably indicate nutrient limitation for all forest ecosystems. We feel both techniques should be corroborated with fertilization trials prior to use on a wider basis.


  1. P. M. Vitousek and R. W. Howarth, “Nitrogen limitation on land and in the sea: how can it occur?,” Biogeochemistry, vol. 13, no. 2, 87 pages, 1991. View at: Publisher Site | Google Scholar
  2. P. M. Vitousek, J. D. Aber, R. W. Howarth et al., “Human alteration of the global nitrogen cycle: sources and consequences,” Ecological Applications, vol. 7, no. 3, 737 pages, 1997. View at: Publisher Site | Google Scholar
  3. W. Koerselman and A. F. M. Meuleman, “The vegetation N:P ratio: a new tool to detect the nature of nutrient limitation,” Journal of Applied Ecology, vol. 33, no. 6, 1441 pages, 1996. View at: Publisher Site | Google Scholar
  4. Z.-Q. Cai and F. Bongers, “Contrasting nitrogen and phosphorus resorption efficiencies in trees and lianas from a tropical montane rain forest in Xishuangbanna, south-west China,” Journal of Tropical Ecology, vol. 23, no. 1, 115 pages, 2007. View at: Publisher Site | Google Scholar
  5. M. E. Fenn, M. A. Poth, and D. W. Johnson, “Evidence for nitrogen saturation in the San Bernardino Mountains in southern California,” Forest Ecology and Management, vol. 82, no. 1–3, 211 pages, 1996. View at: Publisher Site | Google Scholar
  6. A. Gallardo and F. Covelo, “Spatial pattern and scale of leaf N and P concentration in a Quercus robur population,” Plant and Soil, vol. 273, no. 1-2, 269 pages, 2005. View at: Publisher Site | Google Scholar
  7. J. D. May, S. B. Burdette, F. S. Gilliam, and M. B. Adams, “Interspecific divergence in foliar nutrient dynamics and stem growth in a temperate forest in response to chronic nitrogen inputs,” Canadian Journal of Forest Research, vol. 35, no. 5, 1023 pages, 2005. View at: Publisher Site | Google Scholar
  8. E. B. Schilling and B. G. Lockaby, “Relationships between productivity and nutrient circulation within two contrasting southeastern U.S. floodplain forests,” Wetlands, vol. 26, no. 1, 181 pages, 2006. View at: Publisher Site | Google Scholar
  9. J. T. Tessier and D. J. Raynal, “Use of nitrogen to phosphorus ratios in plant tissue as an indicator of nutrient limitation and nitrogen saturation,” Journal of Applied Ecology, vol. 40, no. 3, 523 pages, 2003. View at: Publisher Site | Google Scholar
  10. T. M. Tibbets and M. C. Molles, Jr., “C:N:P stoichiometry of dominant riparian trees and arthropods along the Middle Rio Grande,” Freshwater Biology, vol. 50, no. 11, 1882 pages, 2005. View at: Publisher Site | Google Scholar
  11. T. E. Wood, D. Lawrence, and D. A. Clark, “Variation in leaf litter nutrients of a Costa Rican rain forest is related to precipitation,” Biogeochemistry, vol. 73, no. 2, 417 pages, 2005. View at: Publisher Site | Google Scholar
  12. B. C. Blair and I. Perfecto, “Nutrient content and substrate effect on fine root density and size distribution in a Nicaraguan rain forest,” Biotropica, vol. 33, no. 4, 697 pages, 2001. View at: Publisher Site | Google Scholar
  13. E. Cuevas and E. Medina, “Nutrient dynamics within amazonian forests—II: fine root growth, nutrient availability and leaf litter decomposition,” Oecologia, vol. 76, no. 2, 222 pages, 1988. View at: Publisher Site | Google Scholar
  14. S. K. Gleeson and R. E. Good, “Root allocation and multiple nutrient limitation in the New Jersey Pinelands,” Ecology Letters, vol. 6, no. 3, 220 pages, 2003. View at: Publisher Site | Google Scholar
  15. S. E. Gress, T. D. Nichols, C. C. Northcraft, and W. T. Peterjohn, “Nutrient limitation in soils exhibiting differing nitrogen availabilities: what lies beyond nitrogen saturation?,” Ecology, vol. 88, no. 1, 119 pages, 2007. View at: Publisher Site | Google Scholar
  16. W. T. Peterjohn, C. J. Foster, M. J. Christ, and M. B. Adams, “Patterns of nitrogen availability within a forested watershed exhibiting symptoms of nitrogen saturation,” Forest Ecology and Management, vol. 119, no. 1–3, 247 pages, 1999. View at: Publisher Site | Google Scholar
  17. J. W. Raich, R. H. Riley, and P. M. Vitousek, “Use of root-ingrowth cores to assess nutrient limitations in forest ecosystems,” Canadian Journal of Forest Research, vol. 24, no. 10, 2135 pages, 1994. View at: Publisher Site | Google Scholar
  18. C. G. Stewart, “A test of nutrient limitation in two tropical montane forests using root ingrowth cores,” Biotropica, vol. 32, no. 2, 369 pages, 2000. View at: Publisher Site | Google Scholar
  19. M. do Rosario, G. Oliveira, M. van Noordwijk et al., “Auger sampling, ingrowth cores, and pinboard methods,” in Root Methods: A Handbook, A. L. Smit, A. G. Bengough, C. Engels, M. van Noordwijk, S. Pellerin, and S. C. van de Geijn, Eds., p. 175, Springer, Berlin, Germany, 2000. View at: Google Scholar
  20. B. G. Lockaby and W. H. Conner, “N:P balance in wetland forests: productivity across a biogeochemical continuum,” The Botanical Review, vol. 65, no. 2, 171 pages, 1999. View at: Publisher Site | Google Scholar
  21. C. H. Wharton, W. M. Kitchens, E. C. Pendleton, and T. W. Sipe, “The ecology of bottomland hardwood swamps of the Southeast: a community profile,” FWS/OBS, U.S. Fish and Wildlife Service, Biological Service Program, Washington, DC, USA, 1982. View at: Google Scholar
  22. B. G. Lockaby, F. C. Thornton, R. H. Jones, and R. G. Clawson, “Ecological responses of an oligotrophic floodplain forest to harvesting,” Journal of Environmental Quality, vol. 23, no. 5, 901 pages, 1994. View at: Google Scholar
  23. United States Department of Agriculture, Soil survey of Calhoun and Early Counties, Georgia, U. S. Department of Agriculture, Soil Conservation Service, Washington, DC, USA, 1985.
  24. United States Department of Agriculture, Soil survey of Baker and Mitchell Counties, Georgia, U. S. Department of Agriculture, Soil Conservation Service, Washington, DC, USA, 1986.
  25. A. Mehlich, “Determination of P, K, Na, Ca, Mg and NH4,” Soil Test Division Mimeo, North Carolina Department of Agriculture, Raleigh, NC, USA, 1953. View at: Google Scholar
  26. H. D. Chapman and P. F Pratt, Methods of Analysis for Soils, Plants and Waters, University of California, Division of Agricultural Sciences, Berkely, Calif, USA, 1961.
  27. K. T. Killingbeck, “Nutrients in senesced leaves: keys to the search for potential resorption and resorption proficiency,” Ecology, vol. 77, no. 6, 1716 pages, 1996. View at: Publisher Site | Google Scholar
  28. A. Hodge, D. Robinson, B. S. Griffiths, and A. H. Fitter, “Why plants bother: root proliferation results in increased nitrogen capture from an organic patch when two grasses compete,” Plant, Cell & Environment, vol. 22, no. 7, 811 pages, 1999. View at: Publisher Site | Google Scholar
  29. D. Robinson, “The response of plants to nonuniform supplies of nutrients,” New Phytologist, vol. 127, no. 4, 635 pages, 1994. View at: Publisher Site | Google Scholar
  30. R. Aerts and F. S. Chapin, “The mineral nutrition of wild plants revisited: a re-evaluation of processes and patterns,” Advances in Ecological Research, vol. 30, 1 pages, 2000. View at: Google Scholar
  31. P. M. Vitousek, D. R. Turner, and K. Kitayama, “Foliar nutrients during long-term soil development in Hawaiian montane rain forest,” Ecology, vol. 76, no. 3, 712 pages, 1995. View at: Publisher Site | Google Scholar
  32. S. Güsewell, “N:P ratios in terrestrial plants: variation and functional significance,” New Phytologist, vol. 164, no. 2, 243 pages, 2004. View at: Publisher Site | Google Scholar
  33. H. Olde Venterink, M. J. Wassen, A. W. M. Verkroost, and P. C. de Ruiter, “Species richness-productivity patterns differ between N-, P-, and K-limited wetlands,” Ecology, vol. 84, no. 8, 2191 pages, 2003. View at: Publisher Site | Google Scholar

Copyright © 2008 Matthew A. Neatrour et al. This is an open access article distributed under the Creative Commons Attribution License, which permits unrestricted use, distribution, and reproduction in any medium, provided the original work is properly cited.

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