Piracicaba-SP-Brasil) 2 Universidade Estadual Paulista UNESP Rua Francisco Degni s/n, quitandinha, Araraquara

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1 Scientific registration n : 2277 Symposium n : 1 Presentation : poster On the interaction of Pontoscolex Corethrurus (Müler,1857) and the microbiology of tropical soils Sur l'interaction de Pontoscolex Corethrurus (Müler,1857) et la microbiologie des sols tropicaux BERNARDES Fernando Flávio 1, RIBEIRO Cristina Maria 1, KLEIN Stanlei Ivair 2 1 Engenheiro Agrônomo (Rua Campos Salles, 2186; Bairro Independência; CEP ; Piracicaba-SP-Brasil) minhoca@carpa.ciagri.usp.br 2 Universidade Estadual Paulista UNESP Rua Francisco Degni s/n, quitandinha, Araraquara SP CEP , Brasil stanley@iq.unesp.br INTRODUCTION The Pontoscolex corethrurus is the most abundant species in Brazil, being found in all soils of the country, from the extreme south (Michaelsem, 192; Cordeiro, 191; Vanucci, 195 and Knapper, 1969) to the Amazon (Michaelsem, 1900; Stephenson, 190, Righi, 1967, 1978 and Ayres, 1981). This fact is due to their resistance to dry seasons and their survival strategies. These animals occur in different population densities. In Mexico, they can amount up to 1,70,000 individuals ha -1, consuming 00 tons of soil year -1 (Lavelle 1981). In Brazil they can sum up to 2,0,000 individuals há -1, with annual intake of 1500 tons of soil year -1 (Bernardes and Kiehl, 199). The P. corethrurus reproduce only at temperatures between 2 and 27 ºC, being of low activity in soils with low field capacity of water retention (pf 2.5) (Bernardes and Kiehl, 199). However, they have great capacity to adapt to soils of different ph values, texture and organic matter content (Bernardes et al, 1997), having a demographic growth of type r and a short reproduction cycle, therefore being able to multiply themselves faster than the other types of earthworms. This fact is explained by their ecological strategies (mesohumic endogeic) and their great mutualist digestive efficiency associated to the soil microflora (Lavelle, 1988). The interactions between the micro-organisms and the P. corethrurus during the transit of the soil inside their gut is known (Barois and Lavelle, 1986). The microflora of the soil is also favored by the presence of the earthworm, for it induces the increase of the microbial activity of the original soil (Guerra, 1981). Therefore, it is quite probable the existence of a correlation between the population density and the activity of the P. corethrurus with the activity of micro-organisms in the soil. The present work had the objective of studying the microbial activity of the soils and the activity of P. corethrurus, as a function of time. The influence of two soils of different chemical and physical properties and the addition of organic matter was also investigated. 1

2 METODOLOGY The experiment was conducted under laboratory conditions, under a controlled temperature of 25 ºC and 75% of water retention capacity, for these are the ideal conditions for the survival and reproduction of this species (Bernardes and Kiehl, 199). The treatments consisted of two soils, the oxissols sand (OS) and clay (OC), whose properties can be seen in Tables 1 and 2, and in two levels of organic matter (0 or 18 g cattle manure/ Kg soil). It was used three clitellated individuals per treatment, with three repetitions for each experiment. The soils were incubated during thirteen weeks, with weekly quantification of the following parameters: cocoons, youngsters, casts and CO 2. The separation of the casts was done by the method of sieve separation using 2mm mesh sieves (Bernardes and Ribeiro, 1988), and the microbial activity was determined by the method of Waksman and Starkey (192) of CO 2 evolution. The soils were dried in air and sieved (2mm) to remove any previous macro aggregates. Cattle manure was air dried and sieved (0.5 mm). Each experiment was accompanied by a control where no earthworms were involved. RESULTS AND DISCUSSION In the experiments conducted without the addition of manure, the rates of reproduction of P. corethrurus were higher in OS, but the addition of manure in the complementary experiments levelled the differences. These results must be related to the microbial activity, for even in the treatments without manure the OS soil still presented higher microbial activity than OC. It was observed in the essay that the reproduction of the species was paralysed when the soil respiration fell below mg CO 2 /g soil per day, this value eventually occurring in both types of soils in the absence of manure. In fact, the values in Tables to 6 show that with both types of soil this limiting value is achieved within the rd week of experiments without manure, the OC soils treated with manure being the of soil which kept respiration values above the limit until the 12th week. These measures corroborate the idea that the microbial activity, which represents the feeding of the geophagus earthworm, is profoundly related to the activity of those earthworms. As a fact, in the OS soil with manure which presented the higher rate of earthworm reproduction, the minimum limit of soil respiration was not reached during the time of the experiment (1 weeks). It is important to note that there were performed control experiments without the addition of earthworms, and that our measurements of soil respirometry in those systems showed a statistical equivalence with the values shown in Tables to 6 (experiments involving earthworms). This equivalence asserts the validity of the methodology used for the study of the microbial activity and the activity of the P. corethrurus. It is interesting to note that such observations on the effect of the dietary habits of the geophagus earthworms have been largely negligenced in the international literature; the authors themselves do not have any knowledge of similar work being done by other groups. In the experiments conducted with the addition of cattle manure it was noticed an increase in the number of cocoons and youngsters in both types of soil (Fig. 1). Curiously, it was noticed that in OC the total production of casts also increased as a function of manure (and the population increase), the contrary happening in OS, where the total production of 2

3 casts diminished as a function of the presence of manure (and of the population increase)(fig. 1). If one multiplies the value corresponding to the production of casts per week by the production of CO 2 of the soil before it is consumed, one obtains a value which represents the quantity of microbial biomass contained in the soil which was in fact consumed by the earthworms, measured in mg CO 2. In all treatments of the experiment it was observed that the reproduction was paralysed when this value fell below 6.0 mg CO 2 liberated by the soil to be consumed by the three adult individuals per week (Tables to 6). For this species it seems to be this value which represents the minimum activity of the micro-organisms which must be contained in the soils to be ingested by the individuals to maintain their reproduction capacity. It had been rather curious to note that the division of that constant (6.0 mg CO 2 ) by the weekly values of CO 2 obtained from the soils (values in the 6th column in Tables to 6), lead to a second number, which now represents the minimum quantity of soil that the earthworm should ingest to maintain its reproduction (this theoretical number is shown in the 8th column of the Tables to 6). This means that the lowest the soil respiration, more soil the earthworm should ingest/digest to detonate its reproductive process. We noted that for values below mg CO 2 /g of soil, the quantity of soil that should be consumed by the three individuals is superior to the quantity of soil that the individuals effectively consumed during the experiments, which characterised the condition of impossibility of the earthworm of obtaining enough energy for reproduction (see, for instance, the entries and in Table ). These magic numbers in fact reflect the curious variations in the reproduction of the earthworms: in the studied soil which is considered to be the more fertile, the addition of manure increased the microbial activity to higher extents, which caused a rapid consume of the compostable organic matter, therefore inducing a rapid decay of that activity and consequently, turning difficult in time the activity of the earthworms, which consequently will reproduce less. We therefore conclude that the better the soil, the return of organic matter to the soil should be effected in shorter periods for the geophagus earthworms to maintain an optimal relation to the microbial activity. This last factor is important because when the organic matter stays on the surface, the microbial activity is slow and gradual, and therefore the geophagus earthworms have the time of reproductive activity augmented. We shall note that this ideal condition of interaction between earthworms and micro-organisms is such as that which naturally occurs in some conditions of minimum tillage plantation. The results presented here also show that it should be inefficient to inoculate tropical soils whose values of respirometry are inferior to 0,088 mg CO 2 /g soil with the species studied. Therefore, to increase soil respiration, which corresponds to the microbial activity, it will be necessary the addition of a source of organic matter to produce an effective inoculation. According to our results it was observed that although P. corethrurus have a well developed mutualist strategy (Lavelle, 1988), it still depends on other abiotic factors, as well as of the natural activity of the micro-organisms of the soil before it being ingested by the earthworm. In fact, if only the mutualist strategy was to be operative, than the richest soil should present higher levels of earthworm activity. We found that such a condition was not really met in our controlled conditions; therefore, we suggest that the levels of compostable organic matter in the soil is a primary parameter to be considered regarding geophagous earthworms activity. In this way, we should point out that the rapid

4 humification process which occur with the incorporation of the organic matter in the soil should be deleterious to the action of the earthworms. CONCLUSION This is the first work, to the best of our knowledge, which study the relation of the dietary habits with the activity of tropical geophagus earthworms P. corethrurus. We noted a direct relation of the activity of that earthworm and the microbial activity, in such a way that below a critical value of soil respiration of mg CO 2 /g soil per week, the reproduction of the earthworms ceases completely. That value represents therefore the minimum value of activity of micro-organisms which must be contained in the soil to maintain the reproduction of P. corethrurus. It was also developed a second method, which determinates the quantity of soil (containing micro-organisms) that represents the minimum necessary the earthworm has to consume to maintain the reproduction of the species. The addition of manure to the soil caused an increase in the microbial activity, which favours the reproduction behaviour and the production of casts by the earthworms. These results strongly suggest the intimate relation between the natural activity of the micro-organisms in the soil and the activity of the tropical geophagus earthworms. Therefore, besides the quantisation of the physical and chemical characteristics of soil, our study suggest that the measure of the microbial activity is also necessary for the determination of the viability of the inoculation of that earthworm in new areas, ass well as to estimate its work in time and in space. REFERENCES AYRES, I. & Guera, R.A.T. (1981) Água como fator limitante na distribuição das minhocas da Amazônia Central. Acta Amazônica.11 (1), BAROIS I, & LAVELLE P (1986). Changes in respiration rate and some physicochemical properties of a tropical soil during transit through Pontoscolex corethrurus (Glossoscolecidæ, Oligochæta). Soil Biology and Biochemistry, 18 (5) BERNARDES, F.F.; KIEHL, J.C. (199) Alteração no crescimento e na atividade de minhoca do gênero Pontoscolex pela adição de matéria orgânica e carbonato de cálcio. In: C. B. C. S., 2., Goiânia, 199. Resumos. Goiânia, Sociedade Brasileira de Ciência do Solo, BERNARDES, F.F.; KIEHL, J.C. (199) Comportamento da minhoca Pontoscolex corethrurus sob diferentes condições de temperatura e umidade do solo. In: Reunião Brasileira de Fertilidade do solo e Nutrição de Plantas, 21., Petrolina, 199. Anais. SBCS, BERNARDES, F.F.; RIBEIRO, C.M. and KLEIN, S.K. (1997) Ocorrência de P. corethrurus em diferentes estados brasileiros, e a sua relação com a fertilidade dos solos, Acta Biológica Leopoldencia, in press. BERNARDES F.F; RIBEIRO C.M. (1988) Método para determinação da produção de coprólitos de Pontoscolex corethrurus. To be submitted for publication.

5 CORDEIRO, E. (191) Notas sobre sobre os oligoquetos del Uruguay, An. Mus. Argent. Ciênc. Naturais. 6: p GUERRA, R. T. & ASAKAWA, N. (1981) Efeito da Presença e do Número de Indivíduos de Pontoscolex coretrurus Sobre a população total de Microorganísmos no solo. Acta Amazônica. 11(2): KNÄPPER, C. F. U. Variação anual dos Caractere Genitais de Espécies do Gênero Pheretima (oligochaeta). Rev. Bras. Biologia. 29 () p LAVELLE, P. Earthworm activities and the soil system. Biol. Fert. Soils 6:27-251, LAVELLE, P. Stratégies de reproduction chez les vers de terre. Acta Oecol. 2: p MICHAELSEM, W. Oligochaeta Das tierreich. R. Friedländer & Sohn, 10: p MICHAELSEM, W. Oligochaeten Von Peru und Westpatagonien. Fedd. Goteborgs Mus. 2: p RIGHI, J. Nota sobre as Oligochaetas da Amazônia. Acta Amazônica. 8 (): p RIGHI, J. Oligochaetas da região amazônica. Atas sobre a biota Amazônica.5: p STEPHESON, J. The Oligochaeta. Claredon Press, Oxford, 987p VANUCCI, M. Biological notes: on the glossoscolecidae earthworm Pontoscolex coretrurus. Dusenia. (5): p Waksman SA; Starkey RL (192) Microbiological analysis of soil as an index of fertility. Soil Sci. 17, 11. Key words: earthworms, Pontoscolex, microbiology, respirometry Mots clés : vers de terre, Pontoscolex, microbiologie, respirométrie Table 1. Physical properties of the soils used OC=oxissol clay; OS=oxissol sand Sand Clay Silt OC 0,8 61,22 11, OS 8,5 12,07,02 Table 2. Chemical properties of the soils used. (man= cattle manure) PH M.O. P K Ca Mg H+Al SB T V N-NH N-NO IDENTIF. CaCl2 g dm- mg dm- mmolc dm- % mg kg -1 OC-man.,9 9 9, OC+man., , OS-man., 1 6 0, OS+man., ,

6 Cast 10,00 120,00 100,00 80,00 60,00 _ OS - manure _ OS + manure OC - manure OC + manure Cocoon+young 0,00 20,00 0, weeks Fig 1. Casts and cocoon plus youngsters production in two soils (OS e OC) incubated with Pontoscolex corethrurus. either treated or not with cattle manure during 1 weeks. Table. Casts production and reproduction of P. corethrurus related with the microbial activity in oxissol sand without the addition of manure. mgco 2/ mgco 2 Week adult biomass cast cast/week 1 g solo 2 6/mgCO ,1 16,1 0,16 2, ,67 89, 0,122 10, ,19 70,06 0,088 6, ,70 60,17 0,061, ,19 5,0 0,00 2, ,66 9,78 0,05 1, ,1 6,59 0,02 1, ,57,07 0,029 1, ,00 2,00 0,02 1, ,2 0,2 0,02 0, ,82 8,71 0,022 0, ,21 7,5 0,022 0, ,58 6,12 0,020 0, = quantity of soils ingested by the earthworms, in g of wormcasts per week 2= soil respiration = mg CO 2 per individuals per week = minimum estimated soil consumption for the maintenance of earthworm reproduction 6

7 Table. Casts production and reproduction of P. corethrurus related with the microbial activity in oxissol sand with the addition of manure. CO 2/g solo 2 Week adult biomass cast cast/week 1 6/mgCO ,6 79,6 0,69 50, ,76 55,8 0,9 27, ,07 7,6 0,68 17, ,7, 0,261 11, ,67 0,9 0,250 10, ,98 9, 0,25 9, ,28 8,18 0,200 7, ,58 7,2 0,192 7, ,89 6,65 0,180 6, ,19 6,12 0,177 6, ,50 5,68 0,17 6, ,80 5,2 0,171 6, ,10 5,01 0,170 5,9 5 1,2,, = see legend in Table mgco 2 Table 5. Casts production and reproduction of P. corethrurus related with the microbial activity in oxissol clay without the addition of manure. CO 2/g solo 2 Week adult biomass cast cast/week 1 6/mgCO ,75 92,75 0,125 11, ,28 52,1 0,09, ,82 8,61 0,067 2, ,5 1,8 0,060 1, ,89 27,78 0,056 1, ,2 25,07 0,052 1, ,96 2,1 0,07 1, ,9 21,69 0,0 0, ,0 20,56 0,09 0, ,56 19,66 0,01 0, ,10 18,92 0,02 0, ,6 18,0 0,019 0, ,16 17,78 0,01 0, ,2,,= see legend in Table mgco 2 7

8 Table 5. Casts production and reproduction of P. corethrurus related with the microbial activity in oxissol clay with the addition of manure. CO 2/g solo 2 Week adult biomass cast cast/week 1 6/mgCO ,09 62,09 0,855 5, ,1 65,66 0,61 2, ,5 66,85 0,57 0, ,77 67, 0,02 20, ,00 67,80 0,25 17, ,2 68,0 0,210 1, ,5 68,21 0,170 11, ,68 68, 0,1 9, ,91 68, 0,100 6, ,1 68,51 0,098 6, ,7 68,58 0,092 6, ,60 68,6 0,088 6, ,82 68,68 0,08 5, ,2,,= see legend in Table mgco 2 8