Abstract
Vibrio cholerae, the etiologic agent of cholera, is autochthonous to various aquatic environments. Recently, it was found that chironomid (nonbiting midges) egg masses serve as a reservoir for the cholera bacterium and that flying chironomid adults are possible windborne carriers of V. cholerae non-O1 non-O139. Chironomids are the most widely distributed insect in freshwater. Females deposit egg masses at the water's edge, and each egg mass contains eggs embedded in a gelatinous matrix. Hemagglutinin/protease, an extracellular enzyme of V. cholerae, was found to degrade chironomid egg masses and to prevent them from hatching. In a yearly survey, chironomid populations and the V. cholerae in their egg masses followed phenological succession and interaction of host–pathogen population dynamics. In this report, it is shown via FISH technique that most of the V. cholerae inhabiting the egg mass are in the viable but nonculturable (VBNC) state. The diversity of culturable bacteria from chironomid egg masses collected from two freshwater habitats was determined. In addition to V. cholerae, representatives of the following genera were isolated: Acinetobacter, Aeromonas, Klebsiella, Shewanella, Pseudomonas, Paracoccus, Exiguobacterium, and unidentified bacteria. Three important human pathogens, Aeromonasveronii, A. caviae, and A. hydrophila, were isolated from chironomid egg masses, indicating that chironomid egg masses may be a natural reservoir for pathogenic Aeromonas species in addition to V. cholerae. All isolates of V. cholerae were capable of degrading chironomid egg masses. This may help explain their host–pathogen relationship with chironomids. In contrast, almost none of the other bacteria that were isolated from the egg masses possessed this ability. Studying the interaction between chironomid egg masses, the bacteria inhabiting them, and V. cholerae could contribute to our understanding of the nature of the V. cholerae–egg mass interactions.



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Abbott, SL, Cheung, WKW, Janda, JM (2003) The genus Aeromonas: biochemical characteristics, atypical reactions, and phenotypic identification schemes. J Clin Microbiol 41:2348–2357
Amann, R, Glockner, FO, Neef, A (1997) Modern methods in subsurface microbiology: in situ identification of microorganisms with nucleic acid probe. FEMS Microbiol Rev 20:191–200
Armitage, P, Cranston, PS, Pinder, LCV (1995) The Chironomidae: The Biology and Ecology of Non-Biting Midges. Chapman and Hall, London, UK, p 572
Ashbolt, NJ, Ball, A, Dorsch, M, Turner, C, Cox, P, Chapman, A, Kirov, SM (1995) The identification and human health significance of environmental aeromonads. Water Sci Technol 31:263–269
Banin, E, Israely, T, Kushmaro, A, Loya, Y, Orr, E, Rosenberg, E (2000) Penetration of the coral-bleaching bacterium Vibrio shiloi into Oculina patagonica. Appl Environ Microbiol 66:3031–3036
Bogosian, G, Bourneuf, EV (2001) A matter of bacterial life and death. EMBO Rep 2:770–774
Broza, M, Halpern, M (2001) Chironomids egg masses and Vibrio cholerae. Nature 412:40
Broza, M, Gancz, H, Halpern, M, Kashi, Y (2005) Adult non-biting midges: possible windborne carriers of Vibrio cholerae non-O1 non-O139. Environ Microbiol 7:576–585
Chowdhury, MAR, Yamanaka, H, Miyoshi, SI, Shinoda, S (1990) Ecology and seasonal distribution of Vibrio parahaemolyticus in aquatic environments of a temperate region. FEMS Microb Ecol 74:1–10
Colwell, RR (1996) Global climate and infectious disease: the cholera paradigm. Science 274:2025–2031
Colwell, RR (2000) Viable but nonculturable bacteria: a survival strategy. J Infect Chemother 6:121–125
Colwell, RR, Brayton, PR, Grimes, DJ, Roszak, DB, Huq, SA, Palmer, LM (1985) Viable but non-culturable Vibrio cholerae and related pathogens in the environment: implications for release of genetically engineered microorganisms. BioTechnol 3:817–820
Colwell, R, Huq, A (2001) Marine ecosystems and cholera. Hydrobiologia 460:141–145
Felske, A, Rheims, H, Wolterink, A, Stackebrandt, E, Akkermans, AD (1997) Ribosome analysis reveals prominent activity of an uncultured member of the class Actinobacteria in grassland soils. Microbiol 143:2983–2989
Figueras, MJ, Suarez-Franquet, A, Chacon, MR, Soler, L, Navarro, M, Alejandre, C, Grasa, B, Martinez-Murcia, AJ, Guarro, J (2005) First record of the rare species Aeromonas culicicola from a drinking water supply. Appl Environ Microbiol 71:538–541
Halpern, M, Gancz, H, Broza, M. Kashi, Y (2003) Vibrio cholerae hemagglutinin/protease degrades chironomid egg masses. Appl Environ Microbiol 69:4200–4204
Halpern, M, Broza, YB, Mittler, S, Arakawa, E, Broza, M (2004) Chironomid egg masses as a natural reservoir of Vibrio cholerae non-O1 and non-O139 in freshwater habitats. Microb Ecol 47:341–349
Halpern, M, Raats, D, Lavion, R. Mittler, S (2006) Dependent population dynamics between chironomids (non-biting midges) and Vibrio cholerae. FEMS Microb Ecol 55:98–104
Holmes, P, Niccolls, NH Sartory, DP (1996) The ecology of mesophilic Aeromonas in the aquatic environment. In: Austin, B, Altwegg, M, Gosling, PJ, Joseph, S (Eds.) The Genus Aeromonas. Wiley, Chichester, pp 127–150
Huq, A, Small, EB, West, PA, Huq, MI, Rahman, R, Colwell, RR (1983) Ecological relationships between Vibrio cholerae and planktonic crustacean copepods. Appl Environ Microbiol 45:275–283
Islam, MS, Drasar, BS, Sack, RB (1996) Ecology of Vibrio cholerae: role of aquatic flora and fauna. In: Drasar, BS, Forrest, BD (Eds.) Cholera and the Ecology of Vibrio cholerae. Chapman and Hall, London, UK, pp 187–227
Israely, T, Banin, E, Rosenberg, E (2001) Growth, differentiation and death of Vibrio shiloi in coral tissue as a function of seawater temperature. Aquat Microb Ecol 24:1–8
Lee, KH, Ruby, EG (1995) Symbiotic role of the viable but nonculturable state of Vibrio fischeri in Hawaiian coastal seawater. Appl Environ Microbiol 61:278–283
Long, RA, Rowley, DC, Zamora, E, Liu, J, Bartlett, DH, Azam, F (2005) Antagonistic interactions among marine bacteria impede the proliferation of Vibrio cholerae. Appl Environ Microbiol 71:8531–8536
Munro, PM, Colwell, RR (1996) Fate of Vibrio cholerae O1 in seawater microcosms. Water Res. 30:47–50
Nandi, B, Nandy, RK, Mukhopadhyay, S, Nair, GB, Shimada, T, Ghose, AC (2000) Rapid method for species-specific identification of Vibrio cholerae using primers targeted to the gene of outer membrane protein OmpW. J Clin Microbiol 38:4145–4151
Oliver, JD (1993) Formation of viable but nonculturable cells. In: Kjelleberg, S (Ed.) Starvation in Bacteria. Plenum, New York, pp 239–272
Oliver, JD (2005) The viable but nonculturable state in bacteria. J Microbiol 43:93–100
Oliver, JD, Nilsson, L, Kjelleberg, S (1991) Formation of nonculturable Vibrio vulnificus cells and its relationship to the starvation state. Appl Environ Microbiol 57:2640–2644
Pernthaler, J, Glöckner, O, Unterholzner, S, Alfreider, A, Psenner, R, Amann, R (1998) Seasonal community and population dynamics of pelagic bacteria and archaea in a high mountain lake. Appl Environ Microbiol 64:4299–4306
Pidiyar, V, Kaznowski, A, Narayan, NB, Patole, M, Shouche, YS (2002) Aeromonas culicicola sp. nov., from the midgut of Culex quinquefasciatus. Int J Syst Evol Microbiol 52:1723–1728
Pidiyar, VJ, Jangid, K, Patole, MS, Shouche, YS (2004) Studies on cultured and uncultured microbiota of wild Culex quinquefasciatus mosquito midgut based on 16s ribosomal RNA gene analysis. Am J Trop Med Hyg 70:597–603
Ramaiah, N, Ravel, J, Straube, WL, Hill, RT, Colwell, RR (2002) Entry of Vibrio harveyi and Vibrio fischeri into the viable but nonculturable state. J Appl Microbiol 93:108–116
Richardson, JS, Kiffney, PM (2000) Responses of a macroinvertebrate community from a pristine, southern British Columbia, Canada, stream to metals in experimental mesocosms. Environ Toxicol Chem 19:736–743
Sack, DA, Sack, RB, Nair, GB, Siddique, AK (2004) Cholera. Lancet 363:223–233
Shimada, T, Arakawa, E, Itoh, K, Okitsu, T, Matsushima, A, Asai, Y, Yamai, S, Nakazato, T, Nair, GB, Albert, MJ, Takeda, Y (1994) Extended serotyping scheme for Vibrio cholerae. Curr Microbiol 28:175–178
Winner, RW, Bossel, MW, Farrel, MP (1980) Insect community structure as an index of heavy-metal pollution in lotic ecosystems. Can J Fish Aquat Sci 37:647–655
Ziemke, F, Hofle, MG, Lalucat, J, Rossello-Mora, R (1998) Reclassification of Shewanella putrefaciens Owen's genomic group II as Shewanella baltica sp. nov. Int J Syst Bacteriol 1:179–186
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We thank A. Barbul for his technical assistance with the Confocal Fluorescent microscope.
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Halpern, M., Landsberg, O., Raats, D. et al. Culturable and VBNC Vibrio cholerae: Interactions with Chironomid Egg Masses and Their Bacterial Population. Microb Ecol 53, 285–293 (2007). https://doi.org/10.1007/s00248-006-9094-0
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DOI: https://doi.org/10.1007/s00248-006-9094-0