Bacterial strategies to inhabit acidic environments

Bacteria can inhabit a wide range of environmental conditions, including extremes in pH ranging from 1 to 11. The primary strategy employed by bacteria in acidic environments is to maintain a constant cytoplasmic pH value. However, many data demonstrate that bacteria can grow under conditions in which pH values are out of the range in which cytoplasmic pH is kept constant. Based on these observations, a novel notion was proposed that bacteria have strategies to survive even if the cytoplasm is acidified by low external pH. Under these conditions, bacteria are obliged to use acid-resistant systems, implying that multiple systems having the same physiological role are operating at different cytoplasmic pH values. If this is true, it is quite likely that bacteria have genes that are induced by environmental stimuli under different pH conditions. In fact, acid-inducible genes often respond to another factor(s) besides pH. Furthermore, distinct genes might be required for growth or survival at acid pH under different environmental conditions because functions of many systems are dependent on external conditions. Systems operating at acid pH have been described to date, but numerous genes remain to be identified that function to protect bacteria from an acid challenge. Identification and analysis of these genes is critical, not only to elucidate bacterial physiology, but also to increase the understanding of bacterial pathogenesis.     

Solution structure of an RNA duplex including a C-U base pair

The formation of the C-U base pair in a duplex was observed in solution by means of the temperature profile of (15)N chemical shifts, and the precise geometry of the C-U base pair was also determined by NOE-based structure calculation. From the solution structure of the RNA oligomer, r[CGACUCAGG].r[CCUGCGUCG], it was found that a single C-U mismatch preferred being stacked in the duplex rather than being flipped-out even in solution. Moreover, it adopts an irregular geometry, where the amino nitrogen (N4) of the cytidine and keto-oxygen (O4) of the uridine are within hydrogen-bonding distance, as seen in crystals. To further prove the presence of a hydrogen bond in the C-U pair, we employed a point-labeled cytidine at the exocyclic amino nitrogen of the cytidine in the C-U pair. The temperature profile of its (15)N chemical shift showed a sigmoidal transition curve, indicating the presence of a hydrogen bond in the C-U pair in the duplex.     

Coronatine elicits phytoalexin production in rice leaves (Oryza sativa L.) in the same manner as jasmonic acid

The phytotoxin coronatine induced the accumulation of the flavonoid phytoalexins sakuranetin and momilactone A in rice leaves. Coronatine-inducible sakuranetin production was under the control of kinetin and ascorbic acid (AsA), as observed with jasmonic acid (JA). The effects of kinetin and AsA on the activity of coronatine indicated that coronatine might elicit sakuranetin production in a manner similar to JA. The similarity of both their structures and the manner of elicitation of coronatine and JA suggest that they might interact at the same active site(s) to lead to phytoalexin production.     

Hypoxia relieves X-ray-induced delayed effects in normal human embryo cells

We studied the effect of hypoxia on X-ray-induced delayed effects in normal human embryo cells to elucidate the role of oxidative stress in the susceptibility of cells to induction of genetic instability by radiation. We examined X-ray-induced delayed cell death, giant cell formation, and chromosome aberrations under normally oxygenated (20%) and hypoxic (2%) conditions at 28-38 population doublings postirradiation. The results revealed that hypoxia reduced the X-ray-induced delayed effects, suggesting that radiation enhances cellular oxidative stress, which plays a significant role in determining the susceptibility of irradiated cells to genetic instability. The present study emphasizes the biological significance of epigenetic effects, such as oxygen tension, as well as direct DNA damage in the induction of genetic instability by radiation.     

The primary electron acceptor of green sulfur bacteria,bacteriochlorophyll 663, is chlorophyll a esterified with Δ 2,6-phytadienol

The primary electron acceptor of green sulfur bacteria, bacteriochlorophyll (BChl) 663, was isolated at high purity by an improved purification procedure from a crude reaction center complex, and the molecular structure was determined by fast atom bombardment mass spectroscopy (FAB-mass), ¹H- and ¹³C-NMR spectrometry, double quantum filtered correlation spectroscopy (DQF-COSY), heteronuclear multiple-quantum coherence (HMQC) and heteronuclear multiple-bond correlation (HMBC) spectral measurements. BChl 663 was 2.0 mass units smaller than plant Chl a. The NMR spectra showed that the macrocycle was identical to that of Chl a. In the esterifying alcohol, a singlet P7¹ signal was observed at the high-field side of the singlet P3¹ signal in BChl 663, while a doublet peak of P7¹ overlapped that of P11¹ in Chl a. A signal of P7-proton, seen in Chl a, was lacking, and the P6-proton appeared as a triplet signal near the triplet P2-proton signal in BChl 663. These results indicate the presence in BChl 663 of a C=C double bond between P6 and P7 in addition to that between P2 and P3. The structure of BChl 663 was hence concluded to be Chl a esterified with 2,6-phytadienol instead of phytol. In addition to BChl 663, two molecules of the 13²-epimer of BChl a, BChl a′, were found to be present per reaction center, which may constitute the primary electron donor. This revised version was published online in June 2006 with corrections to the Cover Date.     

Structure of ribulose 1,5-bisphosphate carboxylase/oxygenase gene cluster from a thermophilic hydrogen-oxidizing bacterium, Hydrogenophilus thermoluteolus, and phylogeny of the fructose 1,6-bisphosphate aldolase encoded by cbbA in the cluster

Four genes, cbbO, cbbY, cbbA, and the pyruvate kinase gene (pyk), were found downstream of ribulose 1,5-bisphosphate carboxylase/oxygenase (RubisCO) genes, cbbLS, from a thermophilic hydrogen-oxidizing bacterium, Hydrogenophilus thermoluteolus (formerly Pseudomonas hydrogenothermophila). cbbO was similar to norD in the denitrification gene cluster, and cbbY was similar to cbbY from other autotrophic bacteria. cbbA encoded fructose 1,6-bisphosphate aldolase (FBP aldolase); however, CbbA was little similar to other CbbA proteins. When CbbA was overexpressed in Escherichia coli, overproduction of CbbA was detected by SDS-PAGE. However, the cell extract had slightly higher activity than a cell extract of E. coli without cbbA. Phylogenetic analysis showed class II FBP aldolase divided into classes IIA and IIB, and that CbbA from H. thermoluteolus was in class IIA. Activities of RubisCO and FBP aldolase were examined under autotrophic, mixotrophic, and heterotrophic conditions. The activities of the two enzymes were regulated independently.     

Immunobiological activities of a chemically synthesized lipid A of Porphyromonas gingivalis

A synthetic lipid A of Porphyromonas gingivalis strain 381 (compound PG-381), which is similar to its natural lipid A, demonstrated no or very low endotoxic activities as compared to Escherichia coli-type synthetic lipid A (compound 506). On the other hand, compound PG-381 had stronger hemagglutinating activities on rabbit erythrocytes than compound 506. Compound PG-381 also induced mitogenic responses in spleen cells from lipopolysaccharide (LPS)-hyporesponsive C3H/HeJ mice, as well as LPS-responsive C3H/HeN mice. The addition of polymyxin B resulted in the inhibition of mitogenic activities, however, compound 506 did not show these capacities. Additionally, compound PG-381 showed a lower level of activity in inducing cytokine production in peritoneal macrophages and gingival fibroblasts from C3H/HeN mice, but not C3H/HeJ mice, in comparison to compound 506. Thus, this study demonstrates that the chemical synthesis of lipid A, mimicking the natural lipid A portion of LPS from P. gingivalis, confirms its low endotoxic potency and immunobiological activity.     

Protective effect of antiganglioside antibodies against experimental Trypanosoma brucei infection in mice

Liposome-associated ganglioside antigens (ganglioside GM1 or bovine brain gangliosides) were prepared to facilitate the potential protective efficacy for Trypanosoma brucei. Mice were immunized with liposome-associated ganglioside GM1 or bovine brain gangliosides intraperitoneally (i.p.). After immunization, significantly higher antigen-specific IgG and IgM antibodies were detected in sera than in the nonimmunized control group. When sera from immunized mice were analyzed for isotype distribution, antigen-specific IgG1, IgG2a, and IgG3 antibody responses were also noted. After immunization, mice were challenged i.p. with 1 x 10(2) cells of T. brucei. Sixty percentage of liposome-associated ganglioside GM1-immunized mice survived the infection, and all the mice immunized with bovine brain gangliosides-containing liposomes survived. However, all control mice died within 7 days after infection. These data demonstrate that liposomes containing ganglioside antigens have the potential usefulness for the induction of a protective immune response against T. brucei infection and suggest the possibility of developing vaccines that may ultimately be used for the prevention of trypanosomiasis.     

Further observations on the development of Gongylonema pulchrum in rabbits

Third-stage larvae of Gongylonema pulchrum from naturally infected dung beetles were inoculated orally into 24 rabbits. Worm recovery ranged from 54 to 91% (mean = 67.5%) during the period from 24 hr to 52 wk postinoculation (PI). Two hours PI, the larvae entered the mucosa at the junction of the stomach and esophagus and migrated upward. Early development occurred primarily in pharyngeal mucosa, tongue, and buccal mucosa. The third molt took place 11 days PI and the final molt at 36 days PI. Male worms reached sexual maturity at 7 wk PI and females at 9 wk PI. Adult worms were found mainly in the esophagus but also occurred in the tongue and the wall of the oral cavity after 30 wk PI. Embryonated eggs appeared in the feces of 3 rabbits inoculated with 50 or 100 larvae on days 72-81 PI. Morphologically, the cuticle in young fourth-stage larvae exhibited bosses on the anterior portion on day 11 PI, and the left spicule length : total body length exhibited no remarkable change between 9 and 52 wk PI. The latter finding confirms the utility of the ratio for identification of the nematode.     

Oxygen transport by low and normal oxygen affinity hemoglobin vesicles in extreme hemodilution

The oxygen transport capacity of phospholipid vesicles encapsulating purified Hb (HbV) produced with a Po(2) at which Hb is 50% saturated (P 50 ) of 8 (HbV(8)) and 29 mmHg (HbV(29)) was investigated in the hamster chamber window model by using microvascular measurements to determine oxygen delivery during extreme hemodilution. Two isovolemic hemodilution steps were performed with 5% recombinant albumin (rHSA) until Hct was 35% of baseline. Isovolemic exchange was continued using HbV suspended in rHSA solution to a total [Hb] of 5.7 g/dl in blood. P(50) was modified by coencapsulating pyridoxal 5'-phosphate. Final Hct was 11% for the HbV groups, with a plasma [Hb] of 2.1 +/- 0.1 g/dl after exchange with HbV(8) or HbV(29). A reference group was hemodiluted to Hct 11% with only rHSA. All groups showed stable blood pressure and heart rate. Arterial oxygen tensions were significantly higher than baseline for the HbV groups and the rHSA group and significantly lower for the HbV groups compared with the rHSA group. Blood pressure was significantly higher for the HbV(8) group compared with the HbV(29) group. Arteriolar and venular blood flows were significantly higher than baseline for the HbV groups. Microvascular oxygen delivery and extraction were similar for the HbV groups but lower for the rHSA group (P < 0.05). Venular and tissue Po(2) were statistically higher for the HbV(8) vs. the HbV(29) and rHSA groups (P < 0.05). Improved tissue Po(2) is obtained when red blood cells deliver oxygen in combination with a high- rather than low-affinity oxygen carrier.