Ruthenica, 2014, vol. 24, No. 1: 31-33. © Ruthenica, 2014 Published electronically May 21, 2014. http: www.ruthenica.com

Disease of the shells of Indian apple snails (: globosa)

AJESH K., SREEJITH K.* 1 Department of Biotechnology and Microbiology, Kannur University, Kerala–670 661 India * Corresponding author, E-mail: [email protected]

ABSTRACT. The present investigation was undertak- may pose a threat to some groups of people residing en to study a shell disease of the , Pila within its range. globosa. Observations were made in June-July in four We report the occurrence of a shell disease in consecutive years. The disease first appears as blisters . Initially, blister formation in the peri- in the periostracum and then, once the periostracum is ostracum is seen (Fig. 1B). As the disease progress- lost from these lesions, dissolution of the underlying es, more blisters appear. Once the protein coat has calcified layer. The numerically predominant bacterial genera in the lesions included Aeromonas, Pseudomo- been lost, the calcified layer appears as white patch- nas, Escherichia and Listeria. Communication de- es. This is followed by deterioration of the shell and scribes this previously unreported shell disease, which cavity formation (Fig. 1C) when exposed to envi- may be a health problem in apple snails. ronment factors such as varying pH. Once the pH of the environment drops, the exposed calcium part starts to dissolve. Problems may arise, when holes are formed in the cavity, exposing the soft tissues below. The operculum, which helps to prevent The apple snail, Pila globosa (Swainson, 1822) drying out during aestivation [Meenakshi, 1964] is is a vital component of playing an im- also vulnerable to deterioration (Fig. 1D). portant role in the maintenance of aquatic ecosys- The infected specimens used in this study were tems. They are widely distributed in Nepal and collected from paddy fields near brackish water in south western Asia. Apple snails are exceptionally Kannur district, Kerala, India during June – July of well adapted to tropical regions with periods of 2010, 2011, 2012 and 2013 on the onset of the drought alternated with periods of excessive rainfall monsoon. The study was undertaken with the in- [Subba Rao, Dey, 1989]. Pila globosa occurs in all tention of isolating the causative agent from the types of temporary and permanent water bodies like infected snails. Diseased snails were brought to the ponds, canals and ditches [Jahan et al., 2001]. laboratory and maintained under natural tempera- Though the is widespread, in certain re- ture and moisture regimes. All experimental proce- gions like Nepal, it is decreasing due to habitat loss, dures were carried out under aseptic conditions. and fish poisoning [Subba Rao, Dey, 1989]. Mol- The shell was washed and then surface sterilized luscs are excellent sources of trace and minor with 70% ethanol, carefully without rupturing the elements that are essential for the growth and devel- blisters. Then with the aid of a surgical blade, the opment of humans and they are also used as nutri- blister was cut open and using a micropipette, 5 µL ent supplements for domestic and birds of the blister fluid was transferred to tubes contain- [Baby et al., 2010]. In Bangladesh and some parts ing an enrichment broth (10 ml) prepared in snail of India, Pila globosa is used as a protein supple- infusion (prepared by boiling healthy Pila globosa ment for humans as well as in aquaculture, such as in double distilled water) at pH 7, containing 1% shrimp and catfish farming [Nath et al., 2008; tryptone, 0.5% yeast extract and 1% NaCl. We Wilkins, Lee, 2002]. It has also been tested as a have found this medium to be preferable to other biocontrol agent for the aquatic weed Salvinia mo- bacteriological media used for studying the bacterial lesta [Thomas, 1975]. The natural pathogens of flora of snails. The inoculated tubes were incubated apple snails are not well known, though Godan at room temperature (30±2° C) for 3 days. The [1983] reported the association of microorganisms turbid growth in the liquid medium was then streaked especially protozoa, both as parasites and as sym- onto solidified snail infusion media and incubated at bionts or commensals, but was cautious in assess- room temperature for three days before the colo- ing the role of parasites or pathogens in population nies were counted. Differently looking colonies was regulation. Occurrence of disease in Pila globosa isolated in pure cultures. Three gram negative and 32 K. Ajesh , K. Sreejith

FIG. 1. A. Healthy Pila globosa with undamaged shell. B. Multiple blisters (white arrows) on the shell. C. Shell deterioration and cavity formation. D. Diseased operculum.

РИС. 1. A. Здоровая Pila globosa с неповрежденной раковиной. B. Многочисленные пузыри (отмечены белыми стрелка- ми) на раковине. C. Разрушение раковины и образование полостей. D. Зараженная крышечка.

one gram positive bacteria were frequently isolated to look for the natural pathogens, but also to explore from the lesions of different samples. Direct scan- their role in population control. ning electron microscopy examination of the vesi- References cle fluid also revealed the presence of a few bacilli. The phenotypic characterization of the isolated strains Altwegg M. 1995. Aeromonas and Plesiomonas, In: revealed the genera Aeromonas, Pseudomonas, Es- Murray, P.R., Baron, E.J., Pfaller, M.A., Tenover F.C., Yolken R.H. (eds.) Manual of clinical microbi- cherichia and Listeria, identified based on the de- ology. ASM Press., Washington, DC, pp. 507-516. scriptions of Altwegg [1995], Bille et al. [1995] and Baby R.L., Hassan I., Kabir K.A., Naser M.N. 2010. Holt et al. [2000]. The isolates identified may be Nutrient analysis of some commercially important opportunistic ones taken advantage of the small molluscs of Bangladesh. Journal of Scientific Re- breaks formed on the periostracum. This study search, 2: 390-396. reveals for the first time, the occurrence of this Bille J., Rocourt J., Swaminathan B. 1995. Listeria, Ery- shell disease and the observation made here is obvi- sipelothrix, and Kurthia. In: Murray P.R., Baron E.J., Pfaller M.A., Tenover F.C., Yolken R.H (eds.) ously a health problem in apple snails. The disease Manual of clinical microbiology. ASM Press., may pose a great threat to populations of this re- Washington, DC: 346-352. newable natural resource but more studies are nece- Godan D. 1983. Pest slugs and snails: Biology and ssary to reveal the details of infections including control. Springer-Verlag, Berlin, 445 p. host specificity. The results not only encourage one Holt J.U., Krieg N.R., Sneath P.H.A., Staley J.T., Will- Shell disease in Pila globosa 33

iams, S.T. 2000. Bergey’s manual of determinative Заболевание раковины у индийской ампуллярии bacteriology, Lippincott Williams & Wilkins, Phila- (Ampullariidae: Pila globosa) delphia, USA.

Jahan M. S., Akter M.S., Sarker M.M., Rahaman M.R., * Pramanik M.N. 2001. Growth ecology of Pila glo- АДЖЕШ К., СРИДЖИТ К. bosa (Swainson) (: Pilidae) in stimulat- Department of Biotechnology and Microbiology, Kannur Uni- ed habitat. Pakistan Journal of Biological Scienc- versity, Kerala–670 661 India; * автор для переписки es, 4: 581-584. [email protected] Meenakshi V.R. 1964. Aestivation in the Indian apple snail Pila. 1. Adaptation in natural and experimen- tal conditions. Comparative Biochemistry and Phys- РЕЗЮМЕ. Настоящая работа была проведена для iology, 11: 379-386. изучения заболевания раковины у пресноводного Nath R.D., Rahi M.L., Hossain G.S., Huq K.A. 2008. моллюска Pila globosa. Наблюдения проводились в Bangladesh status of fresh water snail in Khulna июне-июле в течение четырех последовательных лет. district. Bangladesh Research Publication Jour- Заболевание проявляется вначале в появлении пу- nal, 1: 337-347. зырей на периостракуме, а после того как периост- Subba Rao N.V., Dey A. 1989. Freshwater molluscs in ракум исчезает в местах повреждений, в растворе- aquaculture. In: Handbook of Freshwater Mol- нии подлежащих кальцифицированных слоев. До- luscs of India. Zoological Survey of India: 225-232. минирующими по численностями родами бактерий Thomas K.J. 1975. Biological control of Salvinia by the в местах повреждений являются Aeromonas, snail Pila globosa Swainson. Biological Journal Pseudomonas, Escherichia и Listeria. Это первое опи- of Linnean Society, 7: 243-247. сание заболевания, которое моэжет вызывать про- Wickins J.F., Lee, D.O’C. 2002. Crustacean farming: блемы со здоровьем у ампуллярий. Ranching and culture. Blackwell Science, Oxford, 480 p. 