Acid Shock Research Articles

Factors affecting the extreme acid resistance of Escherichia coli O157:H7

When stationary phase Escherichia coli O157:H7 cells were subjected to extreme acid shock (pH 2·0, 6 h, 37°C) cell survival was as great as 10%, but culture conditions greatly affected the acid resistance. Anaerobic cultures were more resistant to extreme acid shock if the glucose concentration of the growth medium was high, acids accumulated, and pH declined. By varying pH and acetate concentration, it was possible to demonstrate a high correlation (R2=0·86) between undissociated acetate and extreme acid resistance. Because dissociated acetate and extreme acid resistance were poorly correlated (R2<0·01), it appeared that the pH effects were being mediated via acetate dissociation. Propionate and butyrate were as effective as acetate, but formate, lactate, benzoate and the uncoupler, carbonylcyanide m -chlorophenylhydrazone (CCCP), were much less effective in promoting extreme acid-resistance. Acetate, propionate, butyrate, benzoate and CCCP all decreased the intracellular pH of E. coli O157:H7, but the correlation between intracellular pH and extreme acid resistance was low (R2<0·01). Cultures grown aerobically only needed half as much acetate to induce extreme acid resistance as those grown anaerobically, and the addition of the reducing agent, cysteine, to anaerobic media made the stationary phase cells less responsive to acetate. An rpoS mutant of E. coli O157:H7 was at least 100-fold more sensitive to acid shock than the wild-type, and large amounts of acetate were needed to promote even a small increase in viability.

Membrane cyclopropane fatty acid content is a major factor in acid resistance of Escherichia coli.

Cyclopropane fatty acid (CFA) formation is a post-synthetic modification of the lipid bilayer that occurs as cultures of Escherichia coli and many other bacteria enter stationary phase. We report the first distinct phenotype for this membrane modification; early stationary phase cultures of strains lacking CFA (as a result of a null mutation in the cfa gene) are abnormally sensitive to killing by a rapid shift from neutral pH to pH 3. This sensitivity to acid shock is dependent on CFA itself because resistance to acid shock is restored to cfa mutant strains by incorporation of CFAs from the growth medium or by introduction of a functional cfa gene on a plasmid. The synthesis of CFA depends in part on the RpoS sigma factor, but the role of RpoS in resistance to acid shock involves additional factors because strains with null mutations in both cfa and rpoS are more sensitive to acid shock than either single mutant strain. Exponential phase cultures of E. coli are much more sensitive to acid shock than stationary phase cultures, but survival is greatly increased if the exponential phase cultures are exposed to moderately acid conditions (pH 5) before shift to pH 3. We show that exposure to moderately acid conditions gives a marked increase in cfa transcription. The efficiency of the survival of acid shock is extremely strain dependent, even among putative wild-type strains. Much, but not all, of this variability can be explained by the partially or totally defective RpoS alleles carried by many strains.

Survival of Escherichia coli O157:H7 in dried beef powder as affected by water activity, sodium chloride content and temperature

A study was carried out to determine the rate of inactivation of Escherichia coli O157:H7 in beef powder as affected by aw(0·34±0·06±0·01), sodium chloride content (0·5, 3·0 and 20%) and temperature (5 and 25° C) over an 8-week storage time. Retention of viability of acid-adapted, acid-shocked, and control cells was determined. Overall, there were no significant differences (P≤0·05) in survival among the three types of cells subjected to the same test parameters, suggesting that mechanisms associated with induction of acid adaptation or acid shock do not result in cross protection against dehydration or osmotic stresses. At each awand within cell type, an increase in sodium chloride concentration resulted in significant reductions in the number of viable cells after a given storage time. Regardless of cell type, survival was significantly higher in beef powder containing 0·5 or 3% sodium chloride compared to powder containing 20% salt. The rate of inactivation was enhanced at aw0·34±0·06 compared to aw0·68±0·01 and at 25° C compared to 5° C.

The response to stationary‐phase stress conditions in Escherichia coli : role and regulation of the glutamic acid decarboxylase system

Inducible bacterial amino acid decarboxylases are expressed at the end of active cell division to counteract acidification of the extracellular environment during fermentative growth. It has been proposed that acid resistance in some enteric bacteria strictly relies on a glutamic acid-dependent system. The Escherichia coli chromosome contains distinct genes encoding two biochemically identical isoforms of glutamic acid decarboxylase, GadA and GadB. The gadC gene, located downstream of gadB, has been proposed to encode a putative antiporter implicated in the export of gamma-aminobutyrate, the glutamic acid decarboxylation product. In the present work, we provide in vivo evidence that gadC is co-transcribed with gadB and that the functional glutamic acid-dependent system requires the activities of both GadA/B and GadC. We also found that expression of gad genes is positively regulated by acidic shock, salt stress and stationary growth phase. Mutations in hns, the gene for the histone-like protein H-NS, cause derepressed expression of the gad genes, whereas the rpoS mutation abrogates gad transcription even in the hns background. According to our results, the master regulators H-NS and RpoS are hierarchically involved in the transcriptional control of gad expression: H-NS prevents gad expression during the exponential growth whereas the alternative sigma factor RpoS relieves H-NS repression during the stationary phase, directly or indirectly accounting for transcription of gad genes.

Acid Adaptation of Listeria monocytogenes Strains Does Not Offer Cross-Protection against an Activated Lactoperoxidase System

Listeria monocytogenes has been implicated in foodborne illness outbreaks involving several types of cheeses made from acidified milk. Acid shock response (ASR) and acid tolerance response (ATR) could be possible reasons for its survival. The ASR and ATR of three strains of L. monocytogenes (V7, V37, and CA) in skim milk acidified to pH 4.0 and 3.5 with lactic acid and held at 32°C were studied. Studies were also done to determine if acid adaptation of the organism enhanced survival in the presence of an activated lactoperoxidase system. The cells were directly shocked at pH 4.0 and 3.5 in skim milk to study the ASR. To study the ATR, cells were initially adapted in skim milk at a mild pH of 5.5 for the equivalent of one generation before being shocked at pH 4.0 and 3.5 in skim milk. Cells adapted at pH 5.5 in tryptic soy broth without dextrose and nonadapted cells were challenged at pH 4.5 in skim milk with or without an activated lactoperoxidase system. In all cases, viability and pH were measured 24 or 48 h after challenge. In pH 4.0 skim milk, for all three strains, the adapted cell population survived better (0.5 to 1.0 log higher) than that of nonadapted cells for 24 h. In pH 3.5 skim milk, the acid-adapted populations of all three strains were 3 to 4 logs greater than those of nonadapted cells at 6 h. The acid adapted cells of all three strains had survival rates similar to those of the nonadapted cells at pH 4.5 both in the presence and absence of an activated lactoperoxidase system. It was also evident that these strains do not exhibit an adaptive ATR at pH 4.5, although they do at lower pH levels (pH 4.0 and 3.5). Survival due to the ATR was better seen at pH 3.5 than at pH 4.0.

Expressed by stress

Enteric bacteria such as Helicobacter pylori, Vibrio cholerae, Salmonella spp. and Escherichia coli survive extreme fluctuations in pH, contributing to their commensal and pathogenic roles. As we enter the genomic age, the identification of co-regulated groups of genes and of the enormous upregulation of particular genes will become routine. These phenomena are interpreted as an indicator of environmental adaptation and, hopefully, will be followed up by biochemical efforts to identify the physiological results of the genetic regulation. Two recent publications have addressed the induction of genes under pH extremes, one taking a genetic approach, the other a proteomic approach. By RNA abundance analysis, Normark’s group1 have found a novel gene, asr, that is highly induced in response to external acid shock and encodes a secreted 11.6-kDa polypeptide. The Asr protein is not essential for growth at neutral or acidic pH and its expression is induced only in response to acid, not to deviations in temperature or osmolarity. The two-component PhoB–PhoR regulatory system probably mediates asr expression; however, the extracytoplasmic sensor for transduction of the environmental signal is not known. In contrast, Blankenhorn and colleagues2 have identified several proteins by 2-dimensional gel electrophoresis and amino-terminal sequencing, which are induced by permeant acid treatment (which lowers the pH of the cytosol) and by severe base treatment. They also investigated whether aerobic or anaerobic conditions affect induction of these proteins. YfiD, a homologue of pyruvate–formate lysase, is induced by internal acidification and is not affected by oxygenation. Induction of another protein, AhpC, occurs during both acid and oxidative stresses. The induction of GadA under alkaline conditions is a surprising finding, as this amino acid decarboxylase is expected to alkalize cells grown under acid conditions. Further biochemical and genetic studies may elucidate mechanisms of crosstalk between virulence factor expression and adaptation to the bacterial environment.

Characterization of Lactobacillus collinoides response to heat, acid and ethanol treatments

Tolerance to stress and cross-protection in Lactobacillus collinoides were examined after exposure to ethanol, acid or heat shock. Ethanol and heat-adapted cells demonstrate induced homologous␣tolerance and cross-resistance to acid stress. No cross-protection of acid-adapted cells against ethanol and heat stresses was observed. Heat was the only pretreatment leading to cross-protection against the two other stresses. Whole-cell protein extract analysis revealed that each treatment induced a battery of stress proteins; the synthesis of some of these polypeptides being induced by more than one condition. The greatest overlap was observed between ethanol and heat treatments. Ten proteins were found to be common to these stresses.

The acid-inducible asr gene in Escherichia coli: transcriptional control by the phoBR operon.

Escherichia coli responds to external acidification (pH 4.0 to 5.0) by synthesizing a newly identified, approximately 450-nucleotide RNA component. At maximal levels of induction it is one of the most abundant small RNAs in the cell and is relatively stable bacterial RNA. The acid-inducible RNA was purified, and the gene encoding it, designated asr (for acid shock RNA), mapped at 35.98 min on the E. coli chromosome. Analysis of the asr DNA sequence revealed an open reading frame coding for a 111-amino-acid polypeptide with a deduced molecular mass of approximately 11.6 kDa. According to computer-assisted analysis, the predicted polypeptide contains a typical signal sequence of 30 amino acids and might represent either a periplasmic or an outer membrane protein. The asr gene cloned downstream from a T7 promoter was translated in vivo after transcription using a T7 RNA polymerase transcription system. Expression of a plasmid-encoded asr::lacZ fusion under a native asr promoter was reduced approximately 15-fold in a complex medium, such as Luria-Bertani medium, versus the minimal medium. Transcription of the chromosomal asr was abolished in the presence of a phoB-phoR (a two-component regulatory system, controlling the pho regulon inducible by phosphate starvation) deletion mutant. Acid-mediated induction of the asr gene in the Delta(phoB-phoR) mutant strain was restored by introduction of the plasmid with cloned phoB-phoR genes. Primer extension analysis of the asr transcript revealed a region similar to the Pho box (the consensus sequence found in promoters transcriptionally activated by the PhoB protein) upstream from the determined transcription start. The asr promoter DNA region was demonstrated to bind PhoB protein in vitro. We discuss our results in terms of how bacteria might employ the phoB-phoR regulatory system to sense an external acidity and regulate transcription of the asr gene.

Streptococcus mutans ffh, a gene encoding a homologue of the 54 kDa subunit of the signal recognition particle, is involved in resistance to acid stress.

The ability of Streptococcus mutans, a bacterial pathogen associated with dental caries, to tolerate rapid drops in plaque pH (acidurance), is considered an important virulence factor. To study this trait, Tn917 mutants of S. mutans strain JH1005 which display acid sensitivity have been isolated and partially characterized. In this paper, the characterization of one of these mutants, AS17, is reported. Preliminary sequence analysis revealed that the transposon insertion in AS17 occurred in the intergenic region of a two-gene locus which has been named sat for secretion and acid tolerance. This locus displays a high degree of homology to the ylxM-ffh operon of Bacillus subtilis. The sat+ locus was cloned by complementation of a conditional Escherichia coli ffh mutant with an S. mutans genomic library. Sequencing of the complementing clone identified the intact ylxM and ffh genes as well as a partial ORF with homology to the proUlopuAC gene of B. subtilis which encodes the binding protein of the ProU/OpuA osmoregulated glycine betaine transport system. RNA dot blot experiments indicated steady-state levels of ffh mRNA in the mutant that were approximately eightfold lower compared to parental levels. This suggests a partial polar effect of the sat-1::Tn917 mutation on ffh expression. Upon acid shock (pH 5), wild-type ffh mRNA levels were found to increase approximately four- to eightfold compared to unstressed (pH 7.5) levels. Mutant levels remained unaltered under the same conditions. Experiments designed to investigate the origins of the acid-sensitivity of the mutant revealed a lack of an acid-adaptive/tolerance response. Assays of proton-extruding ATPase (H+/ATPase) specific activity measured with purified membranes derived from acid-shocked AS17 showed twofold lower levels compared to the parent strain. Also, AS17 was found to be unable to ferment sorbitol although it was able to grow in glucose and a variety of other sugar substrates. These findings suggest that Ffh may be involved in the maintenance of a functional membrane protein composition during adaptation of S. mutans to changing environmental conditions.

Role of external calcium in homeostasis of intracellular pH in the cyanobacterium Anabaena sp. strain PCC7120 exposed to low pH

The role of external Ca2+ in the homeostasis of intracellular pH (pHi) of Anabaena sp. strain PCC7120 in response to a decrease in the external pH (pHex) has been studied in cell suspensions. Increase in cytoplasmic pH after acid shock is dependent on the presence of Ca2+ in the medium. The observed Ca2+ -mediated alkalization of the cytoplasm depends on the extent of the shift in external pH. Acid pH shifts resulted in an increased permeability of the cytoplasmic membrane to protons, which could be reversed by increasing the concentration of Ca2+ in the medium. Thus, the ability of Ca2+ to increase cytoplasmic pH might be correlated with an inhibition of net proton uptake by increasing concentrations of external Ca2+ under these conditions. This combined response resulted in the generation and maintenance of a larger pH gradient (ΔpH) at acid external pH values. All Ca2+ channel blockers tested, such as verapamil and LaCl3 , inhibited the observed Ca2+ -mediated response. On the other hand, the Ca ionophore calcimycin (compound A23187) was agonistic, and stimulated both cytoplasmic alkalization and inhibition of net proton uptake. The protonophorous uncoupler carbonylcyanide m-chlorophenyl hydrazone, inhibited this Ca2+ -mediated response, whereas monensin, an inhibitor of the Na+ /H+ antiporter, had no significant effect. The results of the present study suggest that an influx of Ca2+ from the extracellular space is required for the regulation of cytoplasmic pH in Anabaena sp. strain PCC7120 exposed to low external pH values.

Surface-associated heat shock proteins of Legionella pneumophila and Helicobacter pylori: roles in pathogenesis and immunity.

Bacterial heat shock proteins (Hsps) are abundantly produced during the course of most microbial infections and are often targeted by the mammalian immune response. While Hsps have been well characterized for their roles in protein folding and secretion activities, little attention has been given to their participation in pathogenesis. In the case of Legionella pneumophila, an aquatic intracellular parasite of protozoa and cause of Legionnaires' disease, Hsp60 is uniquely located in the periplasm and on the bacterial surface. Surface-associated Hsp60 promotes attachment and invasion in a HeLa cell model and may alter an early step associated with the fusion of phagosomes with lysosomes. Avirulent strains of L. pneumophila containing defined mutations in several dot/icm genes are defective in localizing Hsp60 onto their surface and are reduced approximately 1000-fold in their invasiveness towards HeLa cells. For the ulcer-causing bacterium Helicobacter pylori, surface-associated Hsp60 and Hsp70 mediate attachment to gastric epithelial cells. The increased expression of these Hsps following acid shock correlates with both increased association with and inflammation of the gastric mucosa. A role for Hsps in colonization, mucosal infection and in promoting inflammation is discussed.

Surface-associated heat shock proteins ofLegionella pneumophila andHelicobacter pylori: Roles in pathogenesis and immunity

Bacterial heat shock proteins (Hsps) are abundantly produced during the course of most microbial infections and are often targeted by the mammalian immune response. While Hsps have been well characterized for their roles in protein folding and secretion activities, little attention has been given to their participation in pathogenesis. In the case of Legionella pneumophila, an aquatic intracellular parasite of protozoa and cause of Legionnaires' disease, Hsp60 is uniquely located in the periplasm and on the bacterial surface. Surface-associated Hsp60 promotes attachment and invasion in a HeLa cell model and may alter an early step associated with the fusion of phagosomes with lysosomes. Avirulent strains of L. pneumophila containing defined mutations in several dot/icm genes are defective in localizing Hsp60 onto their surface and are reduced approximately 1000-fold in their invasiveness towards HeLa cells. For the ulcer-causing bacterium Helicobacter pylori, surface-associated Hsp60 and Hsp70 mediate attachment to gastric epithelial cells. The increased expression of these Hsps following acid shock correlates with both increased association with and inflammation of the gastric mucosa. A role for Hsps in colonization, mucosal infection and in promoting inflammation is discussed. Infect. Dis. Obstet. Gynecol. 7:58–63, 1999. © 1999 Wiley-Liss, Inc.

Isolation and Characterization of Acid- and Bile-Tolerant Isolates from Strains of Lactobacillus acidophilus

Lactic acid bacteria have been reported to be useful as a health adjunct and are commonly added to food as the delivery mechanism. The literature contains many conflicting observations for their proposed benefits, and the mechanism of action is undefined. One source of variation is the large number of strains used without proper controls supplemented. Additionally, many of the organisms are not characterized for their acid shock response or the acid-tolerance response, which are known to vary among bacterial species. Our objective was to isolate acid-resistant and bile-resistant variants of Lactobacillus acidophilus and to determine the phenotypic changes. The acid- and bile-tolerant isolates were obtained using natural selection techniques after sequential exposure to hydrochloric acid (pH 3.5 to 7.0) and mixed bile salts. The acid- and bile-tolerant isolates retained their ability to grow at pH 3.5 with 0.3% bile after the selective pressure was removed and reapplied. Isolates varied from their parents for stability in freezing, lactose utilization, protease activity, aminopeptidase activity, plasmid profile, and cell-wall fatty acid profile. These data suggest that the isolated acid- and bile-tolerant isolates possess growth advantages over that of the parents under stress conditions and may be considered as candidates for probiotic strains after further characterization with animal models.

Influence of acid tolerance responses on survival, growth, and thermal cross-protection of Escherichia coli O157:H7 in acidified media and fruit juices

A study was done to determine survival and growth characteristics of acid-adapted, acid-shocked, and control cells of Escherichia coli O157:H7 inoculated into tryptic soy broth (TSB) acidified with organic acids and three commercial brands of apple cider and orange juice. The three types of cells behaved similarly in TSB acidified with acetic acid; however, in TSB (pH 3.9) acidified with lactic acid, acid-adapted cells were more tolerant than acid-shocked cells which, in turn, were more tolerant than control cells. The ability of the three types of cells to grow after inoculation into acidified TSB, then plated on tryptic soy agar containing sodium chloride was determined. Tolerance of acid-adapted cells and, less markedly, acid-shocked cells to sodium chloride was diminished, compared to control cells. The pathogen showed extraordinary tolerance to the low pH of apple cider and orange juice held at 5 or 25°C for up to 42 days. Growth occurred in one brand of apple cider (pH 3.98) incubated at 25°C. Regardless of test parameters, there was no indication that cell types differed in tolerance to the acidic environment in apple cider or orange juice. Survival of control, acid-adapted, and acid-shocked cells heated in apple cider and orange juice was studied. Within each apple cider or orange juice, D 52°C-values of acid-adapted cells were considerably higher than those of acid-shocked or control cells, which indicates that heat tolerance can be substantially enhanced by acid adaptation compared to acid shock.

Differential Regulation of Parallel Mitogen-Activated Protein Kinases in Cardiac Myocytes Revealed by Phosphatase Inhibition

Previous studies have suggested that the contribution of inducible phosphatases to ERK MAPK deactivation is both cell-type- and agonist-specific. The aim of this study was to define the role of inducible phosphatases in ERK MAPK regulation in cardiac myocytes. We examined the kinetics of activation/deactivation of ERK MAPKs following the exposure of cardiac myocytes to endothelin-1 or phorbol ester. Deactivation was prevented by inhibition of protein synthesis indicating a contribution of inducible phosphatases. In contrast, okadaic acid failed to prolong ERK MAPK activation, but activated three myelin basic protein kinases (MBPKs, 55, 62, and 87 kDa) and two c-Jun kinases (46 and 55 kDa). Although the identity of the MBPKs is unknown, the c-Jun kinases corresponded to JNK MAPKs. Simultaneous exposure of cardiac myocytes to okadaic acid and osmotic shock potentiated JNK MAPK activation. Thus, inducible phosphatases regulate ERK MAPK deactivation, whereas okadaic acid-sensitive phosphatases regulate JNK MAPKs and three novel MBPKs.

Grain feeding and the dissemination of acid-resistant Escherichia coli from cattle.

The gastric stomach of humans is a barrier to food-borne pathogens, but Escherichia coli can survive at pH 2.0 if it is grown under mildly acidic conditions. Cattle are a natural reservoir for pathogenic E. coli, and cattle fed mostly grain had lower colonic pH and more acid-resistant E. coli than cattle fed only hay. On the basis of numbers and survival after acid shock, cattle that were fed grain had 10(6)-fold more acid-resistant E. coli than cattle fed hay, but a brief period of hay feeding decreased the acid-resistant count substantially.

PH-dependent activation of the alternative transcriptional factor σ B in Bacillus subtilis

The expression of the sigB gene of Bacillus subtilis was analysed in response to a mild acid shock. This gene is subject to σ B-dependent regulation. It has been found that the expression of sigB is induced as part of the acid-tolerant response. In that respect sigB is similar to the previously described gene gsiB which is also a member of the σ B regulon. Through this induction, the σ B regulon provides protection against acid shock. Besides its protective role against acid shock, no other general function could be directly associated with the σ B regulon. An acidification of the cytoplasmic environment induces synthesis of general stress proteins in B. subtilis.

PH-dependent activation of the alternative transcriptional factor sigmaB in Bacillus subtilis.

The expression of the sigB gene of Bacillus subtilis was analysed in response to a mild acid shock. This gene is subject to sigmaB-dependent regulation. It has been found that the expression of sigB is induced as part of the acid-tolerant response. In that respect sigB is similar to the previously described gene gsiB which is also a member of the sigmaB regulon. Through this induction, the sigmaB regulon provides protection against acid shock. Besides its protective role against acid shock, no other general function could be directly associated with the sigmaB regulon. An acidification of the cytoplasmic environment induces synthesis of general stress proteins in B. subtilis.

Regulation in the rpoS regulon of Escherichia coli

In Escherichia coli, the transcription factor sigmaS, encoded by rpoS, controls the expression of a large number of genes involved in cellular responses to a diverse number of stresses, including starvation, osmotic stress, acid shock, cold shock, heat shock, oxidative DNA damage, and transition to stationary phase. A list of over 50 genes under the control of rpoS has been compiled. The transcription factor sigmaS acts predominantly as a positive effector, but it does have a negative effect on some genes. The synthesis and accumulation of sigmaS are controlled by mechanisms affecting transcription, translation, proteolysis, and the formation of the holoenzyme complex. Transcriptional control of rpoS involves guanosine 3',5'-bispyrophosphate (ppGpp) and polyphosphate as positive regulators and the cAMP receptor protein - cAMP complex (CRP-cAMP) as a negative regulator. Translation of rpoS mRNA is controlled by a cascade of interacting factors, including Hfq, H-NS, dsrA RNA, LeuO, and oxyS RNA that seem to modulate the stability of a region of secondary structure in the ribosome-binding region of the gene's mRNA. The transcription factor sigmaS is sensitive to proteolysis by ClpPX in a reaction that is promoted by RssB and inhibited by the chaperone DnaK. Despite the demonstrated involvement of so many factors, arguments have been presented suggesting that sensitivity to proteolysis may be the single most important modulator of sigmaS levels. The activity of sigmaS may also be modulated by trehalose and glutamate, which activate holoenzyme formation and promote holoenzyme binding to certain promoters. Key words: transcription, translation, regulation, sigma factor, starvation.

Heat-shock proteins expression in fish central nervous system and its possible relation with water acidosis resistance

The expression of 70 and 60-kDa heat-shock proteins (HSP70 and HSP60) and glial fibrillary acidic protein (GFAP), determined by immunoblotting and immunohistochemical methods, was studied in fish neural tissue; moreover, the possible correlation between the expression of these proteins in neural tissue and fish acidosis resistance was also examined. The HSP and GFAP content was analyzed in four different teleostean fish species (gourami, carp, goldfish and trout) under control conditions and in carp under experimental conditions to induce HSPs expression. Under control conditions, HSP70 and HSP60 expression was similar in gourami, carp and goldfish, but gourami had the highest acidosis resistance; trout had the lowest HSP70 and 60 expression and lowest acidosis resistance. The HSP expression pattern was mainly neuronal under control conditions. HSP expression was induced in carp and the effect of this induction on acidosis resistance was studied. Two methods were used for HSP induction in carp: acid shock (2 h at 4.5 pH) and heat shock (2 h at 33°C). A high acidosis resistance, although non-significant, was observed after heat pretreatment. An important HSP expression was detected in glial cells after induction. GFAP expression showed no association with acidosis resistance under either control or experimental conditions.