Adaptive response to starvation in the fish pathogen Flavobacterium columnare: cell viability and ultrastructural changes

BMC Microbiology

Oscar Olivares-Fuster¹  Wenlong Cai¹  Stacey LaFrentz¹  Covadonga R Arias¹ 

[1] Department of Fisheries and Allied Aquacultures, Aquatic Microbiology Laboratory, Auburn University, 203 Swingle Hall, Auburn, AL, 36849, USA

关键词: Columnaris disease;    Survival;    Ultrastructure;    Starvation;    Flavobacterium columnare;   

Others  :  1145033
DOI  :  10.1186/1471-2180-12-266

 received in 2012-06-26, accepted in 2012-10-31,  发布年份 2012

【 摘 要 】

Background

The ecology of columnaris disease, caused by Flavobacterium columnare, is poorly understood despite the economic losses that this disease inflicts on aquaculture farms worldwide. Currently, the natural reservoir for this pathogen is unknown but limited data have shown its ability to survive in water for extended periods of time. The objective of this study was to describe the ultrastructural changes that F. columnare cells undergo under starvation conditions. Four genetically distinct strains of this pathogen were monitored for 14 days in media without nutrients. Culturability and cell viability was assessed throughout the study. In addition, cell morphology and ultrastructure was analyzed using light microscopy, scanning electron microscopy, and transmission electron microscopy. Revival of starved cells under different nutrient conditions and the virulence potential of the starved cells were also investigated.

Results

Starvation induced unique and consistent morphological changes in all strains studied. Cells maintained their length and did not transition into a shortened, coccus shape as observed in many other Gram negative bacteria. Flavobacterium columnare cells modified their shape by morphing into coiled forms that comprised more than 80% of all the cells after 2 weeks of starvation. Coiled cells remained culturable as determined by using a dilution to extinction strategy. Statistically significant differences in cell viability were found between strains although all were able to survive in absence of nutrients for at least 14 days. In later stages of starvation, an extracellular matrix was observed covering the coiled cells. A difference in growth curves between fresh and starved cultures was evident when cultures were 3-months old but not when cultures were starved for only 1 month. Revival of starved cultures under different nutrients revealed that cells return back to their original elongated rod shape upon encountering nutrients. Challenge experiments shown that starved cells were avirulent for a fish host model.

Conclusions

Specific morphological and ultrastructural changes allowed F. columnare cells to remain viable under adverse conditions. Those changes were reversed by the addition of nutrients. This bacterium can survive in water without nutrients for extended periods of time although long-term starvation appears to decrease cell fitness and resulted in loss of virulence.

【 授权许可】

   
2012 Arias et al.; licensee BioMed Central Ltd.

【 图 表 】

Figure 1.

Figure 2.

Figure 3.

Figure 4.

Figure 5.

Figure 6.

Figure 7.

【 参考文献 】

• [1]Austin B, Austin DA: Bacterial fish pathogens: disease of farmed and wild fish. New York, NY: Springer; 1999.
• [2]Wagner BA, Wise DJ, Khoo LH, Terhune JS: The epidemiology of bacterial diseases in food-size channel catfish. J Aquat Anim Heal 2002, 14:263-272.
• [3]Figueiredo HCP, Klesius PH, Arias CR, Evans J, Shoemaker CA, Pereira DJ, Peixoto MTD: Isolation and characterization of strains of Flavobacterium columnare from Brazil. J Fish Dis 2005, 28(4):199-204.
• [4]Amin NE, Abdallah IS, Faisal M, Easa ME, Alaway T, Alyan SA: Columnaris infection among cultured Nile tilapia Oreochromis niloticus. Antonie Van Leeuwenhoek J Microbiol 1988, 54(6):509-520.
• [5]Decostere A, Haesebrouck F, Van Driessche E, Charlier G, Ducatelle R: Characterization of the adhesion of Flavobacterium columnare (Flexibacter columnaris) to gill tissue. J Fish Dis 1999, 22:465-474.
• [6]Suomalainen LR, Tiirola M, Valtonen ET: Chondroitin AC lyase activity is related to virulence of fish pathogenic Flavobacterium columnare. J Fish Dis 2006, 29:757-763.
• [7]Welker TL, Shoemaker CA, Arias CR, Klesius PH: Transmission and detection of Flavobacterium columnare in channel catfish Ictalurus punctatus. Dis Aquat Org 2005, 63:129-138.
• [8]Fijan NN: The survival of Chondrococcus columnaris in waters of different quality. Bull Off Int Epizoot 1968, 69:1158-1166.
• [9]Chowdhury MBR, Wakabayashi H: Effects of sodium, potassium, calcium and magnesium Ions on the surivival of Flexibacter columnaris in water. Fish Pathol 1988, 23(4):231-235.
• [10]Kunttu HMT, Valtonen ET, Jokinen EI, Suomalainen L-R: Saprophytism of a fish pathogen as a transmission strategy. Epidemics 2009, 1:96-100.
• [11]Poindexter JS: Oligotrophy: fast and famine existence. Adv Microb Ecol 1981, 5:63-89.
• [12]Kjellerberg S, Humphrey BA, Marshall KC: Initial phases of starvation and activity of bacteria at surfaces. Appl Environ Microbiol 1983, 46:978-984.
• [13]Suzina NE, Mulyukin AL, Kozlova AN, Shorokhova AP, Dmitriev VV, Barinova ES, Mokhova ON, El’-Registan GI, Duda VI: Ultrastructure of resting cells of some non-spore forming bacteria. Microbiology 2004, 73:435-447.
• [14]Vatsos IN, Thompson KD, Adams A: Starvation of Flavobacterium psychrophilum in broth, stream water and distilled water. Dis Aquat Org 2003, 56:115-126.
• [15]Arias CR, Welker TL, Shoemaker CA, Abernathy JW, Klesius PH: Genetic fingerprinting of Flavobacterium columnare isolates from cultured fish. J Appl Microbiol 2004, 97:421-428.
• [16]Triyanto K, Wakabayashi H: Genotypic diversity of strains of Flavobacterium columnare from diseased fishes. Fish Pathol 1999, 34:65-71.
• [17]Shoemaker CA, Olivares-Fuster O, Arias CR, Klesius PH: Flavobacterium columnare genomovar influences mortality in channel catfish (Ictalurus punctatus). Vet Microbiol 2008, 127:353-359.
• [18]Shoemaker CA, Arias CR, Klesius PH, Welker TL: Technique for identifying Flavobacterium columnare using whole-cell fatty acid profiles. J Aquat Anim Heal 2005, 17:267-274.
• [19]Arias CR, Cai W, Peatman E, Bullard SA: Catfish hybrid Ictalurus punctatus x I. furcatus exhibit higher resistance to columnaris disease than the parental species. Dis Aquat Org 2012, 100:77-81.
• [20]Thoesen JC: Suggested procedures for the detection and identification of certain finfish and shellfish pathogens. Bethesda, ML: American Fisheries Society-Fish Health Section; 2004.
• [21]Kjelleberg S, Humphrey BA, Marshall KC: Initial phases of starvation and activity of bacteria at surfaces. Appl Environ Microbiol 1983, 46:978-984.
• [22]Wai SN, Mizunoe Y, Yoshida S: How Vibrio cholerae survive during starvation. FEMS Microbiol Lett 1999, 180:123-131.
• [23]Garnjobst L: Cytophaga columnaris (Davis) in pure culture: a myxobacterium pathogenic for fish. J Bacteriol 1945, 49:113-128.
• [24]Bernardet JF, Bowman JP: The genus Flavobacterium. In The Prokaryotes, vol. 7. 3rd edition. Edited by Dworkin M, Falkow S, Rosemberg E, Schleifer K-H, Stackerbrant E. New York: Springer; 2006:481-531.
• [25]Madetoja J, Nystedt S, Wiklund T: Survival and virulence of Flavobacterium psychrophilum in water microcosmoms. FEMS Microbiol Ecol 2003, 43:217-223.
• [26]Moller JD, Barnes AC, Dalsgaard I, Ellis AE: Characterisation of surface blebbing and membrane vesicles produced by Flavobacterium psychrophilum. Dis Aquat Org 2005, 64:201-209.
• [27]Chaiyanan S, Chaiyanan S, Grim C, Maugel T, Huq A, Colwell RR: Ultrastructure of coccoid viable but non-culturable Vibrio cholerae. Environ Microbiol 2007, 9:393-402.
• [28]Mulyukin AL, Suzina NE, Duda VI, El’-Registan GI: Structural and physiological diversity among cystlike resting cells of bacteria of the genus Pseudomonas. Microbiology 2008, 77:455-465.
• [29]Tekedar HC, Karsi A, Gillaspy AF, Dyer DW, Benton NR, Zaitshik J, Vamenta S, Banes MM, Gulsoy N, Aboko-Cole M, et al.: Genome sequence of the fish pathogen Flavobacterium columnare ATCC 49512. J Bacteriol 2012, 194:2763-2764.