Abstract Two Azospirillum brasilense strains, CDJA and A40, capable of growing and producing plant growth-promoting (PGP) substances at the sub-optimal temperature (SOT) of 22°C, were tested for their ability to survive, colonize and enhance wheat growth and yield under field conditions upon inoculation. The response was compared with that of A. brasilense strain, A9, impaired in growth and PGP activities at SOT (22°C) but otherwise comparable to CDJA and A40 at 37°C. A field experiment was carried out in a split-plot design with four levels of N as main plots and three strains and an uninoculated control as subplots. A differential response in the establishment of the strains and in plant growth and yield was obtained, due to the categories of strains, particularly at lower levels of N (0 kg and 40 kg N ha –1 ). The results clearly demonstrated that strains capable of growing and producing PGP substances at SOT are better inocula for wheat. Keywords Azospirillum · Temperature · Inoculation response Introduction Azospirillum inoculation of plants has been tested world- wide in view of their contribution to plant productivity by their ability to fix N 2 , their phytohormone production, siderophore production, enhancement of plant mineral uptake, and NO 3 – production (Bashan and Holguin 1997). Despite many successful experiments using Azospirillum spp. as plant inocula, their commercial application on a large scale has lagged behind, and the main obstacle to their use is the unpredictability and inconsistency of field results (Okon and Labandera-Gonzalez 1994). The results of field experiments with winter crops are more inconsis- tent than those with crops grown under a high tempera- ture or tropical conditions (Sumner 1990; Wani 1990; Pandey et al. 1998), as an optimum temperature (e.g. 37°C) is required for growth and the production of bene- ficial plant growth-promoting (PGP) substances (Tripathi and Klingmuller 1992). Azospirillum survives and estab- lishes poorly in the rhizosphere of crops grown in winter due to the prevailing low temperature (18–25°C) (Harris et al. 1989; Wani 1990; Pandey et al. 1998; Kaushik et al. 2001). The low or non-significant effect of Azospirillum inoculation in winter crops discourages the large-scale use of Azospirillum as a biofertilizer for these as compared to its use in summer crops. In earlier studies, we identified two Azospirillum brasilense strains capable of growing and stimulating plant growth at a sub-optimal temperature (SOT) in microcosms (Kaushik et al. 2001). Their establishment and ability to improve wheat growth were tested using Tn5-lacZ insertion mutants, otherwise isogenic to wild type strains under glasshouse conditions (Kaushik et al. 2000). In the present study these Azospirillum strains were used to study the survival and colonization of inocula in the wheat endorhizosphere in the field, in order to test the hypothesis that the poor effects of Azospirillum are due to the poor survival of inocula in winter. Materials and methods Bacterial cultures and growth media Two A. brasilense strains, CDJA and A40, capable of growing and producing PGP substances at SOT (22°C) and one strain, A9, sensitive to SOT but otherwise efficient at the optimum temperature of 37°C, were obtained from the culture collections of the Division of Microbiology, Indian Agricultural Research Institute (IARI), New Delhi. Their intrinsic antibiotic resistant patterns are given in Table 1. These strains were grown and maintained on N-containing sodium malate medium (Bulow and Dobereiner 1975). A.K. Saxena ( ✉ ) · K.V.B.R. Tilak Division of Microbiology, Indian Agricultural Research Institute, New Delhi, 110012 India e-mail: rajeev_ensc@hotmail.com Tel.: +91-11-5787649 R. Kaushik Division of Environmental Sciences, Indian Agricultural Research Institute, New Delhi, 110012 India Biol Fertil Soils (2002) 35:92–95 DOI 10.1007/s00374-002-0444-x ORIGINAL PAPER R. Kaushik · A.K. Saxena · K.V.B.R. Tilak Can Azospirillum strains capable of growing at a sub-optimal temperature perform better in field-grown-wheat rhizosphere Received: 19 March 2001 / Published online: 14 February 2002 © Springer-Verlag 2002