6.3.1 Pinus
The genusPinusis one of the largest and most important among the coniferous
genera. Pines are widely distributed and mostly found in the Northern Hemisphere.
They range from Alaska to Nicaragua, from Scandinavia to North Africa, and from
Siberia to Sumatra (Krugman and Jenkinson 1974 ). The most commonPinus
sp. found in western North America is lodgepole pine. It is a commercially
important gymnosperm species that grows throughout the Rocky Mountain and
Pacific Coast regions. It extends from Yukon Territory, Canada in the north to Baja
California, Mexico in the south and from the Pacific Ocean in the west to South
Dakota, USA in the east (Lotan and Critchfield 1990 ). Thefirst evidence of
endophytic colonization by plant growth-promoting bacteria in lodgepole pine was
detected by Shishido et al. ( 1995 ). They isolated a bacterial endophyte (strain Pw-2)
from root tissues of lodgepole pine seedlings (<3 years old) naturally regenerating
at a site near Williams Lake, BC, Canada (52°N, 122°W). Preliminary characteri-
zation revealed that strain Pw-2 belongs toBacillus polymyxa(now known as
Paenibacillus polymyxa). The beneficial effects ofB. polymyxaPw-2 were assessed
by re-inoculating it into lodgepole pine and growing in a greenhouse for 9 weeks.
Inoculation with Pw-2 significantly increased shoot height, shoot dry mass, and root
dry mass of lodgepole pine seedlings as compared to the uninoculated controls.
A rifamycin-resistant strain, Pw-2R, was derived from Pw-2 so as to check internal
root colonization of lodgepole pine after re-inoculation (Shishido et al. 1995 ).
Pw-2R endophytically colonized the internal root tissues of lodgepole pine with a
population size in the range of 10
5
cfu/g fresh tissue, 4 weeks after inoculation. In a
subsequent study, Shishido et al. (1996a) ruled out the theory about the involve-
ment of mycorrhizal fungi in growth promotion observed in Pw-2 (or Pw-2R)
inoculated lodgepole pine seedlings. According to theirfindings, Pw-2R enhanced
lodgepole pine seedling biomass significantly (up to 18%) through a mechanism
that is unrelated to mycorrhizal fungi. It was also reported that strain Pw-2 is
involved in elevating the levels of PGP hormones like indole-3-acetic acid
(IAA) and dihydrozeatin riboside (DHZR; a form of cytokinin) produced in
lodgepole pine roots (Bent et al. 2001 ).
In an effort to look for diazotrophic bacterial endophytes in stem and needle
tissues of lodgepole pine trees (>20 years old) and seedlings (2–4 years old)
growing in nutrient-poor (N-limited) forest sites of British Columbia, Canada; Bal
et al. ( 2012 ) isolated an endophytic strain P2b-2R that was capable of growing on
N-free medium (combined carbon medium; Rennie 1981 ) and consistently reduced
significant amounts of acetylene in acetylene reduction assay (ARA) used for
measuring N-fixing activity. Strain P2b-2R (GU132543) was identified as
belonging toP. polymyxa(Bal et al. 2012 ). Since ARA is an indirect method of
measuring the amount of Nfixed, Bal and Chanway (2012a) used a more accurate
method,^15 N isotope dilution method (Danso 1995 ), to assess the amount of N
fixed. In two separate growth trials, P2b-2R inoculated lodgepole pine seedlings
were able to derive 30 and 66% of N directly from the atmosphere 27 and 35 weeks
116 A. Puri et al.