Bacteriological taint of musculuss and digestive piece of land contents of Oreochromis sp. and Labeo sp. reared in a pool supplied with domestic sewerage was enumerated followed by finding of opposition of thermotolerent coliforms for antibiotics. Numbers of bacteriums in musculuss and digestive piece of land contents of fish reflected their densenesss in H2O. Muscles of both the fish species contained high Numberss of entire coliforms ( TC ) and faecal coliforms ( FC ) . Escherichia Coli ( E.coli ) were ne’er recovered from the musculuss but from the digestive piece of land contents of the fish. Ranking of the sum and faecal coliform taint degrees showed a lessening in the order digestive piece of land contents & gt ; musculuss ( P & lt ; .05 ) . The random thermotolerent coliform isolates from these fish variety meats showed opposition in diminishing order for Achromycin ( 82 % ) and Principen ( 65 % ) . 58.11 % ( 68 isolates out of 117 ) of the entire isolates were immune to both Principen and Achromycin. This observation indicates important happening of bacterial population in variety meats of sewerage fed fish with high incidence of antibiotic opposition which may present hazard to angle zoologies and public wellness.
Keywords: Domestic sewerage ; Fish ; Fecal coliforms ; Escherichia coli ; Water quality ; Antibacterial opposition ; Public wellness.
* Corresponding Author, E-mail- samirbancu @ gmail.com
# Both the writers have equal weightage
All around the universe, people both in rural and urban countries have been utilizing domestic wastes to fertilise fish pools ( Strauss et al. 2000 ) . In the bulk of instances, domestic sewerages are applied untreated or merely partly treated through storage ( Strauss 2000 ) . Domestic effluent, rich in foods, used in aquaculture supports the growing of plankton and other micro-organisms which are consumed by the fish with small consumption of other supplemented provender. Recycling of domestic sewerage through aquaculture is an effectual signifier of pollution control, which contributes to be recovery and provides a beginning of low cost animate being protein production.
Domestic sewerage transports a assortment of human pathogenic micro-organisms which may pollute fish flesh when fish is grown in pools having waste H2O ( Niewolak and Tucholski 2000 ) . Besides, municipal effluent consists of immense sums of incompletely metabolized antimicrobic drugs which can take to the development of antibiotic immune bacterium every bit good as immune plasmids ( Wiggins et Al. 1999 ) . The conspicuously affected bacteriums are members of Enterobacteriaceaes and related gm negative rods ( Kelch and Lee 1978 ) . One of the of import concern of effluent piscaries is the taint of fishes by faecal coliforms ( Fapohunda, MacMillan, Marshall and Waites 1994 ) . Their presence in fish intended for human ingestion may represent a possible danger non merely by doing disease but besides because of the possible transportation of antibiotic opposition from aquatic bacteriums to human-infecting bacteriums from nonaquatic beginnings ( Olayemi, Adedayo and Ojo 1991 ) . Therefore, periodic and comprehensive healthful study of effluent piscary is required. For old ages, the group of faecal ( besides called thermotolerant ) coliforms ( FC ) has been the most widely used as faecal taint as their excreted burden is similar or larger than that of infective beings, and their survival clip in the environment longer than that of excreted bacteriums and viruses ( Strauss 1997 ) .
In the present survey, an effort has been made to find the bacteriological taint of musculuss and digestive piece of land contents of Oreochromis sp. and Labeo sp. reared in effluent fed pool. Resistance to two really common antibiotics for random thermotolerent coliform isolates from musculus and digestive piece of land contents of both the fish was besides determined.
Materials and Methods
A sewerage fed pool of Bandipur, Rahara, North 24 Parganas, ( 22A°44’N Latitude and 88°24’E Longitude ) was taken into consideration for this survey and to analyze bacterial burden of H2O and fish. Natural sewerage was wholly of domestic beginning, coming from Titagarh town of North 24 Parganas, West Bengal.
Sampling and Dissection
Fish samples were caught with a net and were instantly transferred to the research lab in containers with pool H2O. They were dissected harmonizing to Buras et Al. 1987. Muscles and digestive piece of land contents were isolated and placed in unfertile glass vass. The tissues were weighed under unfertile conditions, land in a howitzer and suspended in Na chloride ( NaCl ) physiological solution ( 10 milliliter of the solution for each 1 g of the musculus or digestive piece of land content ) . The suspensions were homogenized utilizing Universal Laboratory Aid Type MPW-309 homogenizer, at 1000 revolutions per minute, for 10 proceedingss. The homogenates were so serially diluted ( 10-1 to 10-6 for musculuss and 10-1 to 10-7 for digestive piece of land contents ) and inoculated into civilization media. Time slowdown from fish aggregation to the analyses did non transcend 6 hours. Water from sewage-supplied pool was sampled and analysed at the same time with fish sampling. Samples were collected monthly from July 2009 to September 2009.
Lauryl Tryptose ( LT ) Broth at 350C for 48 hour was used for three-tube most-probable-number ( MPN ) presumptive findings of coliforms ( APHA 1998 ) . From all positive presumptive tubings, entire coliforms were confirmed by the formation of gas in any sum in the Durham agitation tubings of superb green milk sugar bile stock ( BGLB ) for 48 hour at 350C.
Faecal Coliforms and E.coli
All positive Lauryl Tryptose ( LT ) MPN tubes to tubes of Escherichia coli ( EC ) Broth followed by incubation at 44.50C for 48 hours constitute a positive faecal coliform trial. The growing from positive EC tubing was so streaked onto Levine Eosin Methylene Blue ( EMB ) Agar home bases and incubated at 35A°C for 18 to 24 h. Colonies from EMB Agar plates typical of E. coli were transferred to Nutrient agar ( NA ) angles from which GIMViC trials were performed where “ G ” -medium is the secondary EC stock, “ I ” -medium is Tryptone stock, “ M ” – and “ V ” -medium is Buffered Glucose stock, and “ C ” -medium is Simmon ‘s Citrate agar. MPN of E. coli was so computed based on the figure of tubings found to incorporate isolates that produce GIMViC reaction patterns characteristic of E. coli ( APHA 2001 ) .
Representatives of typical thermotolerent coliform isolates from fish samples were selected indiscriminately by settlement morphology on Eosin methylene bluish agar and were streaked aseptically several times on newly prepared alimentary agar home bases to obtain pure isolates ( Ogbonna, Sokari and Amaku 2008 ) . Alimentary agar home bases were so supplemented with Principen ( 50Aµg ml-1 ) and tetracycline ( 25Aµgml-1 ) and were used to measure antibiotic susceptibleness forms of 117 pure isolates ( Miranda and Zemelman 2001 ) . 32 isolates from musculuss and 24 isolates from digestive piece of land contents of Oreochromis sp. and 39 strains from musculuss and 22 isolates from digestive piece of land contents of Labeo sp. were subjected to antibiotics sensitivity trial.
Meanss and standard mistakes ( SE ) were calculated. T trial was performed between bacterial concentration of musculuss and digestive piece of land contents of both the fish. A significance degree of 5 % was considered ( Zar 2007 ) .
Table 1: Faecal index bacterial burden ( average + SE, n =3 ) in H2O of waste fed fish pool
( MPN* 100 ml-1 )
( MPN* 100 ml-1 )
( MPN* 100 ml-1 )
*MPN =Most Probable Number
Table 2: Faecal index bacterial burden ( average + SE, n =15, values holding same superior are significantly different at 5 % degree ) in musculuss and digestive piece of land contents of fish collected instantly after crop from wastewater-fed fish pool
Type of Tissue
( MPN 100gm-1 )
( MPN 100gm-1 )
( MPN 100gm-1 )
& lt ; 3
& lt ; 3
Digestive piece of land contents
Bacterial tonss in musculuss and digestive piece of land contents of Oreochromis sp. and Labeo sp. were exceptionally high. Entire coliforms and faecal coliforms were normally found in all analysed fish tissues. Escherichia Coli were non found in the musculuss of either fish. Additionally, nevertheless, thermotolerant Escherichia coli were present in the digestive piece of land contents of both the fish ( Table 2 ) . Bacterial tonss in the fish were significantly higher ( p & lt ; 0.05 ) in the digestive piece of land contents than in comestible musculuss.
Figure 1: Antibacterial opposition of thermotolerent coliforms isolated from fish of sewage-fed pool.
Figure 2: Form of opposition of thermotolerent coliforms to both Principen and Achromycin isolated from musculuss and digestive piece of land contents of Labeo sp. and Oreochromis sp.
Antibiotic opposition form
Of the 117 thermotolerent coliform isolates examined for antibiotic sensitiveness, 82 % ( 96 isolates out of 117 ) were tetracycline resistant and 65 % ( 76 isolates out of 117 ) were ampicillin resistant. 58.11 % ( 68 isolates out of 117 ) of the entire isolates were immune to both antibiotics where as 31 % ( 36 isolates out of 117 ) were immune to individual antibiotic. From the individual antibiotic resistant isolates, 24 % were tetracycline resistant and 7 % were ampicillin immune ( Figure 1 ) . 56.41 % isolates ( 22 isolates out of 39 ) from flesh and 22.72 % ( 5 isolates out of 22 ) from digestive piece of land contents of Labeo sp. showed opposition to both antibiotics whereas 71.87 % isolates from flesh ( 23 isolates out of 32 ) and 75 % ( 18 isolates out of 24 ) from digestive piece of land contents of Oreochromis sp. showed opposition to both Principen and Achromycin ( Figure 2 ) .
Comparison of faecal coliform counts of H2O of Bandipur sewerage fed fish pool with WHO ( World Health Organization ) H2O quality standards ( WHO 1989 ) suggests considerable taint of the first. Bacterial vegetation of fish reflects the bacteriological quality of the H2O from where the fish harvested ( Geldrich and Clarke 1966 ) . Strong correlativity between the bacterial species present in the pool H2O and the fish regardless of the type of fish were besides reported by Buras et Al. 1987 ; Ogbondeminu 1993 ; Apun, Yusofand and Jugang 1999. Therefore, in our survey, entire coliforms, faecal coliforms and E.coli recovered from musculuss and digestive piece of land contents of Oreochromis sp. and Labeo sp. may reflect bacteriological H2O quality of the Bandipur sewerage fed pool.
Faecal coliforms in fish musculuss were recovered when values of FC in H2O were 3.86+3.63×105 MPN 100 ml-1 which were much higher than those recommended by WHO ( 1989 ) in its wellness guidelines on effluent usage in aquaculture. Faecal coliforms in fish reflect the degree of pollution of their environment, as the normal vegetations of fish do non include coliforms ( Cohen and Shuval 1973 ) . Presence of faecal coliforms indicates the presence of faecal stuff from warm-blooded animate beings. However, thermotolerent coliforms include the genera of faecal every bit good as non faecal beginning. E. coli is a species of faecal coliform bacterium that is specific to faecal stuff from worlds and other warm-blooded animate beings ( Bhatia 2008 ) . Environmental Protection Agency ( 1992 ) therefore recommends E. coli as the best faecal index of wellness hazard from H2O. No noticeable incursion of E.coli in musculuss of either fish was found at 1.34+0.95×104 MPN 100 ml-1 of E.coli concentration in H2O of sewerage fed pool ( Table 1 ) . Therefore, the fish flesh qualities at harvest were good on the footing of their E. coli counts. Safety safeguards during fish processing are still needed to avoid cross-contamination due to high accretion of micro-organisms in the digestive piece of land of fish.
In this survey significantly higher Numberss of bacteriums ( P & lt ; 0.05 ) were found in digestive piece of land contents than in musculuss of both of the fishes which may traverse contaminate fish filet. Similar findings were reported by Ogbondeminu and Okoye 1992 ; Fatal et Al. 1993. Bacterial ( entire coliforms, faecal coliforms and E.coli ) loads in digestive piece of land contents of both Oreochromis sp. and Labeo sp. were found to be higher than environing H2O. Same findings were proposed by Ogbondeminu and Okoye ( 1992 ) ; Fatal et Al. ( 1993 ) . High concentrations of bacteriums in digestive piece of land contents are chiefly due to high concentrations of phagocytic cells localized in the bowels of these fish ( Ellis, Munroe and Roberts 1976 ) . These phagocytic cells constitute the first barrier to foreign beings occupying the fish.
A broad scope of thermotolerent coliforms isolated from sewerage fed fish showed opposition to both Principen and Achromycin. Multiple anitibiotic immune fecal coliforms have been observed in effluent across the universe ( Gallert et al. 2005 ) . Antibiotic opposition among random bacterial isolates from different variety meats of fish captured from fecally contaminated H2O with a full scope of opposition ( 00-100 % ) to different common antibiotics of curative and contraceptive usage among human existences and in assorted carnal farms and fish farms was reported by several writers. ( Rhodes et al. 2000 ; Miranda and Zemelman 2001 ; Pathak and Gopal 2005 ) . Thus the beginning of the job of antibiotic opposition bacteriums in effluent pool of Bandipur was fecally contaminated H2O.
Among thermotolerent coliforms recovered from fish, opposition to ampicillin and tetracycline was found in 65 % and 82 % of the isolates, severally. Happening of thermotolerent coliforms with high opposition to ampicillin and tetracycline reflect human influence in the environment ( Andersen and Sandaa 1994 ) . Domestic sewerage enters into the pool environment of the sewerage fed farm with immense antibiotics which are used as medical specialties, as growing boosters or as preventive care and may hold established a selective force per unit area due to a slow debasement of disinfectants favoring farther growing of antibiotic-resistant bacteriums ( Petersen and Dalsgaard 2003 ) . It may possible that these antibiotic resistant bacteriums from effluent may reassign their antibiotic resistant determiners to autochthonal vegetations of fish, arousing their spread and prevalence in aquatic environment.
In the present survey bacteriums resistant to both Principen and Achromycin from digestive piece of land contents were higher in Oreochromis sp. than in Labeo sp. It may be related to detritus feeding wont of Oreochromis sp. by which it is more open to wastes every bit good as disinfectants than Labeo sp. which is a column feeder. Similar findings were reported by Miranda and Zemelman ( 2001 ) with demersal and oceanic fish. Antibiotic immune faecal bacteriums form domestic sewerage may alter nutritionally good enteric microflora with unexpected effects on fish wellness.
Our survey indicates that fish flesh qualities were satisfactory in footings of E.coli counts. In malice of that flesh of both fish showed high Numberss of antibiotic immune thermotolerent coliforms which may include Klebsiella spp. , Citrobactor spp. and Enterobacter spp. ( non faecal beginning ) but till have immense ecological and public wellness deductions specially if the opposition is plasmid mediated so there could be a job associated with the transportation of opposition determiners to human pathogenic bacteriums which may come in in human population through fish ingestion. Harmonizing to Walia et Al. ( 2004 ) antibiotic opposition cistrons against Principen, streptomycin, and Achromycin are known to be movable to other bacteriums.
Therefore, we can state that Effluents and fishes reside there are powerful beginning of antibiotic-resistant bacteriums, which in bend may reassign their opposition cistrons to nonresistant bacteriums ( Schwartz et al. 2003 ) . Several surveies indicate that the environmental conditions in effluent may heighten the likeliness of cistron transportation ( Pote et al. 2003 ) . Mach and Grimes ( 1982 ) demonstrated the high transportation frequences of enteral bacteriums in a effluent. Additionally immune bacteriums may present a hazard of curative jobs to public wellness and fish population. So the survey demands an luxuriant probe on the members of prevailing multidrug immune bacterial microflora associated with sewerage fed fishery along with their plasmids profile as an grounds of connubial transportation of antibiotic opposition cistrons in homo and animate being nutrient concatenation through fish ingestion.