IndiJn Journal or Experi mentJ I Biology Vol. 4 1. March 2003. pp. 248-254

Chitin degrading potential of bacteria from extreme and moderate environment

N N Nawani & B P Kapadnis*

Department of Microbiology. Uni versi ty or Pune. Pune 4 11 007. India

Recei ved 22 No velllber 2002; revised 19 j allllclI ] 2003

Five hundred chitin-degrad ing bacteria were iso lated rrom 20 different locations. Hi gh percentage of potent chitin­degraders was obtained from po lluted regions. Potent chit in- degrading bacteri a were selected by primary and ,eeondary scree ning. Among the se lected isolates. 78% we re rcprcsc ntcd by the gcn us STreplUlllyces. Majority of thc iso lat es had good chitino lys is relati ve to the growt h although iso lates with beller growt h were al so seen. Such isolates arc important 1'0 1' the producti on of SCP from chitinous wastes. The potent iso lates be longed to the genera STrepf()/.'lyces. KirasalUsporia.

Sacc/wwpolyspol'{[. Nocardioides. Nocardiol ls is. I-Ierbidospora. MicrOIllOIlOSP0/'{f. Micwbispo/'{f. I ICTill oplalle ..... SerraTia.

iJacilllis and Pse lidolllollas. Thi s study form s J comprehensive base for the study of divcrsity of chitinolytic syste ms of bacte ri J.

Ch itin , the ~-I , 4 linked polymer of N-acetylgluco­samine is the second most abundant polysaccharide in nature. Chitin is a structural component of the ce ll wa ll s of fungi as well as of shell s or cuticles of arthro­pods, crustaceans, insects and mollusks. The major contri buti on of chi tin to soil is in the form of animal biomass. Similarly, in the marine environment more than lO ll metric tons of chitin is annually produced l. Soil and aquatic systems both act as major harbors of chitin degraders. The biological conversion of chitin by means of microbial chitinases is mainly responsible for the repleni shment of carbon and nitrogen to the atmos­phere , thereby maintaining the ecological balance. Due to multiple app lications of chitinases in biocontrol, waste management, medicine and biotechnology, they become interesting enzymes fo r study.

The present study was aimed to iso late chitin­degrad ing bacteria and understand the diversity of their chitinolytic sys tems. For thi s purpose, soil and water samples were obtained from various locat ions, both ex treme and moderate. considering the fact that it is not necessary to go to the ex treme environments to encounter exoti c diversit/. This study primarily gives an idea of the ecologica l effects on chitin degra­dat ion att ribute of bacteria. A detailed comparison of the chitinolyti c systems, keeping in view the habitat of the iso lates can further reveal some interest ing eco­logical effects.

*Correspondent author: Tel: 9 ! 0205690643 Fax:9 ! 0205690087 Email : bpkapadn is@yahoo.com

Materials and Methods The locati ons of sa mpling included habitats classi­

fied as ex treme and moderate as shown in Table I, fro m where so i I and water samples were co ll ected using poi nt-integrating samplers and by the proce­dures recommended in APHA:l . Hand-held scoops and shovels were used for sampling near-surface so il s.

Isolation and enllllleration o/chiTin degraders Isolation of chitin degraders was done on co ll oidal

chitin agar (CCAt Iso lation was performed by serial dilutions of the samples and plating on buffered) CCA. The number of iso lates capab le of chitin degra­dation was enumerated at three pH values (4.0. 7.0 and 9.5) and isolates hav ing good zone of clearance on CCA were preserved and subjected to primary and secondary screening fo r selection of potent isolates.

Primary screening of chitin degraders Primary screening was performed by spot inoc ul at­

ing the iso lates on CCA buffered to pH 4.0, 7.0 and 9.5 to class ify them into groups based on max imum chitinolys is at ac idic, neutral or alkaline pH. Zone of chitin hydrolysis and colony diameter were recorded up to 7 days at 40°C. Whether the iso lates have a similar range of pH for growth as fo r chit in degrada­tion was determined by monitoring their growth in nutri ent broth or Bennett 's medium at pH 4.0, 7.0 and 9.5 .

Secondary screening 0/ chiTin degraders Secondary screening was performed to check the

ab ility of iso lates to degrade practical grade chitin

.,

NAWAN I & KAPADNIS : CHITI N DEG RADING BACTERI A FROM EXTREME AN D MODERATE EN VIRONMENT 249

T:.Jb lc I - Ci :Jssifi c:J tion of s:J mpli ng locati ons and occurrcncc of chi tinolytic bactcria

Extrcmc locati ons* Po ll utcdt

Modcratc locati on_s __ ..-___ _ Non'-pollu tcdtf

Sali nc Lake, Lonar

Acid soil s, Sana , Ycmcn

Ac id so il s, Ranchi , India

Sassoon Docks, MUlllbai

Mu la river, Punc

Hudayduh, Saud i Arab ia

Mecca, Saudi Arabia

Adcn, Yemcn

Hot springs, Vraj rcshwari , Mum b:l i

Compost, Pune

Mu tha ri ve r, Punc

P:.Jvana ri ver, Punc

Pash:J n Lakc, Punc

Jcddah, S:Judi Arabia

Pcacock Bay, Pune Agri cultu ra l land, Punc Kalewad i, Punc

Compost, Mumba i Laundry Spots, Punc

Potcnt ch itinolyti c b:Jctcri a fo und in -*Ex trcme locat ions: Sl replolllyces, Baci llus, MicroIIlOll o.lp om, Microbispora tPolluted locat ions: SlreplOlllyces, Bacillus, Pseudolllullas, Serr(l{ ia, Micl'OlIIOIIOspom. Nucardiopsis, KilasalU ­

sporia, ACl ill op/oll es

tt Non-polluted loc:J ti ons: Sl rep lUlllyces, Herbiduspom , Noca rd iu iiles, Sacclwl'Opo/yspora

flakes and to produce ex trace llul ar chitinase. 0. 1 ml cell or spore suspension of 00600 0.1 was inoculated in SO ml of chitin medium at optimum pH value and was incubated in a rotary incubator shaker (I SO rpm) at 40°C for 6 days. Ex tracellul ar chitinolytic acti vity in the culture mediu m was determined using swollen chitin as the substrate as described by Monreal and Reese6 at optimum pH of chitinolys is as determined by primary screening. One unit of chitinase ac ti vity was defin ed as the amount of enzy me required to re­lease one flm ol of Gl cNAc in I min under the above­menti oned conditi ons of enzy me reacti on.

Identification oj chitinolytic bacteria The identi fication of ac tinomycetes and other bac­

teri a was based on the methods recommended in the Bergey's Manu al of Sys tematic Bacteri olog/ includ­ing other reco mmended references and Internati onal Streptomyces Projects. The chemotaxonomi c studies were carri ed out fo r actinomycetes onl y by detennina­tion of cell wa ll amino ac id9 and whole cell sugars. Probabili stic identification matri ces were used fo r identification of Streptomyces spec ies based on physiological and biochemi cal charac teri sti cs lO and other genera were manuall y identified.

Results

Isolation and enumeration oJchitin deg raders The advantage of simultaneous iso lation at three

pH va lues th an isolati on on medium of single p I-I va lue was th at thi s method gave the probable relati ve abundance of bac teria capable of degrading chitin in the ac idi c and alkaline pH range. It could be clearl y elucidated th at pH has an effect on the predominance of a paJ1icuiar group of organi sms relati ve to the habitat.

The samples from Lonar Lake with pH in the alkal ine range ex hibited the predominance of iso lates capable of degrading chitin in the alk aline range. Soil samples from agricultural lands at Pune, Mecca, Hudaydah, and Sana had more number of ch itin degraders in compari son to the so il samples from Lonar, Vrajreshwari hot springs and laundry spots. Whereas, an intermedi ate count was seen for soi I sam­ples fro m Aden, Jeddah and Pashan and lowest cou nts were for so il samples from Peacock Bay. In general, the number of chitin degraders in water samples was lesser than for so il samples, where culti vated and garden soil s had a higher number of chitinolyti c bac teri a. The samples with acidic pH like those from Sana and Ranchi had more number of iso lates capab le of degrading chitin in the ac idic pH range. Most of the iso lates in samples with neutral pH like those from Mula, Mutha, Mecca and Jeddah etc. displ ayed more number of bacteri a degrading chitin in the neutral p H range. Thus, pH as a selection fac tor may not be so lely restri cted to chitin degradation but may also be ex tended to other acti viti es of the isolates. Five hu n­dred isolates di sp lay ing good chi ti nolys is were pre­served fo r screening and subsequent selec ti on of po­tent chitin degraders.

Primary screening oj chitin degraders Primary screening enabled the class ification of"

isolates as those exhibiting max imum chitinolys is at acidic, neutral or alkaline pH values . Most of the iso lates with acidic or alkaline pH optima for growth had better chitinolys is at ac idic or alkaline p H respec­tively. The isolates with optimum growth at neutral pH although did not necessarily ex hibit maximum chitin degradation at neutral pH but some of them had better degradability at ac idic pH. Similarly some

250 I DIAN J EXP SIOl, MARCH 2003

isolates w ith optimal growth in the acidic range ex hibited max imum chi tin olys is in neutral range and some w ith optimal grow th in alkaline range had maximum chitinolys is ar pH 7.0. Thus, there were some variati ons in the optimum pH range for growth and chitinolys is. The gross colony morphology of iso­lates could identify maj ority of them as act inomy­cetes; however, iso lates with better chitin degradat ion at alkaline pH were mainly other bacteria .

The iso lates we re further di vi ded into groups based on the rati o o f zone of chitino lys is to co lony diameter. The percent frequencies o f iso lates w ith rati o o f zone of c learance to co lony diameter fall ing in the ranges 1.0- 1.5, 1.5-2.0, 2.0-2.5, 2.5 -3.0, 3.0-3.5, 3.5 -4.0 and 4.0-4.5 'vvere determined. Fig . I di splays the percent freq uency of the iso lates relat ive to the above­mentioned classes. It ca n be seen that most o f the iso­lates from the three ca tegori es (ac idi c, neutral and alka line p H) have rat io in the range 1.5-2.0 and 2.0-2.5. The selec tion of iso lates for secondary screening was done b'1.~ed on rati o o f zone of chitino lys is to col­ony diameter of 2.0 or >2 .0. The iso lates w ith rati o or

zone of chiti nolys is to co lony di ameter ::::: 1.5 w ith a good co lony diameter indicating good grow th were al so se lec ted. Th e larger co lony diameter but margi nal ch il ino lys is indica te either a low ex tracellular chiti­nase act i vity or an intracellular or cc ll bound ehitinase act i vity or rapid utili za ti on o f end-products o f chitin degradation . Non-streptomycete iso lates w i th good

60 ,

I 50 I

if) 40 ~ ro (5 if)

1.5 2.0 2.5 3.0 3.5 4.0 4.5

Ra ti o of zone of ch itin hyd rol ys is to colony diameter

I:ig. 1- !'1'CqUCIlCY dist ribu ti oll o r d ilTcI'C lll groups 01' cilitilloly t ic orgallisms based Oil ratio o r c il itillo lys is to colo llY d iamctcr (1) 1-1

--lO D Ji ll 7.0 - . Ji ll 9.5 0)

chitino lysis were also preferred due to th e unabated capab ility of streptomycetes to degrade chitin. Iso lates w ith nearly simil ar activ iti es at the three pH va lues were al so selec ted for secondary screen ing. Thu s, 200 isolates were selected for secondary sc reening which enabled the selec ti on of strai ns for the stud y of di ver­sity of their ch itinascs.

Secolldwy screenillg oi chitill deg raders The iso lates w i th potent chitinoly ti c <tcti vity were

se lected for secondary sc reening, a graphica l repre­

sentati on of which is gi ven in the Fig. 2. It ca n be

seen that a higher percentage of potent degraders was

obtained from moderate locat ions, although the per­centage of potent chitin degraders from ex treme loca­

ti ons was good relati ve to the number of ex treme

habitats undertaken for the study. Of the moderate

hab itats, higher percentage of good isol ates was from

polluted reg ions than non-pol luted ones. Polluted sites

showed nearl y similar abundance of potelll degraders

amon gst different habitats, whereas in case of non­

polluted habitats, Peacock Bay g~l ve the least efficient

degraders. Polluted reg ions can lead to better adapta­

bi lity of th e ll1 icroorgani sms to their surroundings

besides offering the competi t ion for survival.

Secondary screen i ng was done usi ng pract ical grade chitin fl akes inste: tcl of co lloidal chitin whi ch is

an eas il y access ible substrate as compared to the chi ­tin fl akes. It was seen that higher percentage of bac te­

riJ di sp layed chitinase acti v ity in the ra nge 0-0. i U

Il1r t, whereas streptomycetes had ci1it inase act ivity above 0.2 U Il1r t coverin g 78 % of the isolates. In sev­

cral iso lates ch iti nase acti Vl ty dras ticall y decreased

w ith increase in incubation peri od accompan ied by a decrease in spec ifi c acti v it y although the pro tein con­ten t had either increased or was nearl y rile sa me. Th is could have been due to the degradation of ch itinase either due to protease or that the enzyme producti on is

repressed.

Iso lates (6 1.2%) had biomass protein in the range of 10- 15 mg and 11 .6% iso lates had biomass prote in above I S mg; most o f these iso lates were those w ith a greater co lony size but (mll'gina l chiti nolys is. Such iso lates can be very useful in producti on o f SCP from chi ti nous was tes and thus as a source of pro tein or animal feed. Sirepio/llyces sp. i"ormed the ll1aj or group o f potent chitin degrader:.; whereas non-s trepto­mycetes and other bacteria compr ised a relatively sll1all proporti on. The iso lates w ilh greater chitinase

NAWANI & KAPA DNI S: CHIT IN DEG RA DING BACTER IA FROM EXTREM E AND MODERATE ENVIRON M E T 25 1

(lc ti vity (I ndio I' greater spec ifi c act ivity of chiti nase or a hi gher ce ll protein were selected for furth er studi es. 49 iso lates were finall y selected to study the di versity ex hibited by their chit inolytic systems whi ch were identified to the genus leve l fo r a comparati ve study of their chitinolytic systems.

Occllrrence oj chitin deg rading bacteria The forty-nine iso lates selected for further studies

were identified to the genus level. They were assigned genotypes based on their chemotaxonomic , growth , physiological and biochemi ca l characteri stics as per the class ifi cation desc ri bed in the Bergey's Manual of Systematic Bacteriolog/ and Bergey 's Manual of

Moderate ~

Non-Polluted

38%

Non-Po lluted

Khadak- Huday-was ala dah

Agricul- 4% 15%

Non­Extreme

69%

~~r~ /\ ~ ~ Jeddah

LJ~ S-J 15%

Kale-wadV 1\ V, 15% U Aden

Mecca 13%

17%

Determinati ve Bacteriology' I. 36 isolates belonged to the genu s Streptomyces with a cell wa ll hav ing gly­cine and L-DAP and a fa tty acid profile with satu­rated, iso and anti -i so fatty ac ids. On the bas is of growth , ph ys iolog ica l and biochemical characteri stics, few isolates were found to be close to Streptomyces lIlacrosporlls, St replolllyces lJyg roscopicus, Strepto­lIlyces jUl1lonjinensis, Streptomyces toruloslIs, Strep­tOlllyces heterol1lol'phus, Streptomyces jllipinensis, Streptolllyces cellulosae, St reptomyces roclJ ei. Strep­tomyces halstedii, Streptomyces e4oliatlls. Strepto­l1Iyces albus, St reptol1lyces g ralllinojaciens and St rep­tOlllyces xant/lOcli rolllogenes. However, spec ies were not assigned to the isolates since it was essential to

~ Extreme

Compos t

6%

Acid Saline

Soils lake

35% 46%

Hot Springs

13%

Po lluted

Laundry Pas han 10% 15%

Pavana 1\ \l ~ Sasoon 17% ~~ Docks \lD 13% Muth~ 22% Mula

23%

Fi g. 2 - Habilal- wise di slri buli on of e ffi c ient chilin deg rade rs selec led fo r secondary sc ree ning

252 I 'DIAN J EXl' BIOL, MARCH 2003

conf irm the same using molecular phylogeny approaches. Of the forty-nine potent chitin-degrad ing bacteri a, the genus Slreplolllyces was found to be pre­sent in almost every hab itat. L onar Lake, M ecca and agri cultural land had maximum percentage of potent chitinolytic streptomyce tes, whereas none of these were obta ined from Sassoon Docks and Pashan Lake. Besides streptomycetes, Lonar Lake was inhabited by chitin degrad ing genera of Bacil/lls and MiCrDIl IOIIO­

SPOJ'(l . 'fhe iso late c lass ified as Microll lO ll ospom was char, cteri zed by si ngle non-mot i Ie sess i Ie spores and ce ll wa ll containing amino ac ids IIl eso-DAP, glyc ine and xy lose and arabinose as who le ce ll sugars. The isolates from ac id so il s and hot springs were of genus SlreplOlllyces however, Microbisporrt sp., wh ich has been recen tl y reponed fo r its chitino ly ti c potenti al, was isolated in thi s stu dy from th e hot springsl 2.

Of the polluted habitats, fJacil/lIs sp. was iso lated from Sassoon Docks and strain s o r the genera Psell ­

dOlllOllas , MicrDll lO llOSpom, NOc(lU/iops is and Ser m­

li(l were obtained from Mula, Mutha an d Pavana ri v­ers respectively. The genus Noca rdiops is was charac­terized by the presence of IIl eso-DAP in the ce l l wa ll and no diagnost ic sugars in vvh ole ce ll hyd rolysa tcs. Its substrate mycel ium was dcnsely branched and showed rragmentat ion in later growth phases. Bes ides streptomyce tes, the genera KiwsolOsporia and Ac­

lill op/oll es were ['ound in th e locati ons or laundry. Kitosalo.I'{Jorio had L-DAP, lIleso-DAP, g lyc ine in ce ll wa ll s and ga lactose in whole ce ll hydrolysates whereas Au illop/oll es showcd mati Ie spores in spherical to globose sporangia w ith IIl eso- D A P and glyc ine in ce ll wall and xy lose and arab inose in whole ce ll hydrolysates. Of the non-po lluted locati ons, non­streptomycc te isol ates li ke /-/erbidospom, Noco rdi­

oides and Sacc/wropolyspoJ'(l were round at Huday­dah, Aden and kddah respec t i ve l y . Sacc/lClropoly ­spo /'{/ was characteri zed by the w h i te aer ial mycc l i um and a ce ll wa ll wi th IIl cso-DAP, galactose and arab i­nose. The iso late of the genlls Nocordioides had ce ll wa ll wit h L-DAP and glyci ne and its substrate myce­l iu m showed fragmen tati on and th at o f the genus f-1erbido.lpom had a ce ll wal l w ith lIleso- D A P and who le cell sugar ,1S madurose.

Discussion Soi l is a ve ry ri ch source or organ isms, where se­

lec ti on pressure leads to the enri chment o f a parti cular group or organi sms and in competiti on these groups are L:X pected to ex press ccrtai n phenotypes, wh ich confe r upon thelll an advantage ove r o thers. Chitin is

abundantly found in so il and water, the recyc ling of wh ich is essential for an ecologica l ba lance. Chitin degradat ion is mediated in so il and water by several types o f microorgani sms, of wh ich act inomyce tes playa major role. M ost studi es on chit inases indicate that these enzy mes generall y ex hibit maxim um acti v­ity at pH ranging from 5.0 to 7.0 13 like the chitinase from the well-studied bacterium, Ser rat ia 1/1ClrCeSCeIlS,

which produces chitinase w ith optimu m activity near pH 6.0 and w ith stab ility in a wide pH rangel -I. How­

ever, chitinases w ith pH optima of 4.0 have been found in streptomycetes5 and Microbispom Sp. 12 and

those with pH optima in the alkal ine range have been reported for Streptolllyces albido.flavlIsl 5 and Nocar-

I · . 1/ I Ii d B '11 17 ( IOPS1.\' a JIlS an aCI liS sp. A predominance of chitin degraders in moderate

regions than in ex treme locati ons could be due to their deviat ion from the normal ones in terms of pH and temperature so they allow the survi va l o f limited or­gani sms capab le of thri ving under these conditi ons. T he second reason could be a relati ve ly lesser avail­ab ility of chitin in these environmcnts. This was how­ever, not seen for ac id soil s from Sana, where a good number of chitin degraders were found . Most or the ac tinoillycetes can tolerate pH up to 5.0 <lI1d bL:ing cu lti vab le reg ion, the nutrient availab ili ty o f the so il at Sana must be highcr. Besides, an ac idi c p H can lead to pro l i l'eration of fungi that would i nd i rect i y contri b­ute to the chitin content o r the soil.

In general, a higher predomi nance of bacteri a de­grad ing chi tin at neutral pH was seen in most o r the samples. A n except ion to th ese, were the so il and wa­ter samples obta ined from ex treme type of locat ions li ke sa li ne lakes, ac id so il s and hot sp ri ngs. Iso lation performed at a single p H va lue l im its thc poss ibility or iso lati ng chitin degraders c;lpable of express ing thi s att r ibu te at oth er p H values wh ich might o f reI' en­zy Illes w i th grea ter stab i I i ti cs and thu s o f industrial importance. The resu lts indica te occurrence or chitin degraders in almost every hab i tat stud ied, suggestive of the importance of these iso lates in chitin minerali­zati on anci recyc ling. It is ev ident that acid soils have predominance of chitin degraders ac ti ve at acidic pH and the same is tru e fo r alkaline and neutra l habi tats. Al though, it could be eva lua1.ed that neu trophil es do ex ist in acidic and alka line hab itats, but how far do they contri bute in chitin recyc ling in th e~c habitats is a matter of in trospect ion ill si lil. The relative abun­dance, in general, provides a clue to undcrstanding the el'feet of nutrients and envii'Onlllental fac tors on a par­ticular phenotype.

NAWA 1& KAPADN IS : CHITI N DEGRADING BACTERIA FROM EXTREME AN D MODERATE ENVIRONME T 253

Ac idophiles appear to be confined to acid so il s and litters and neutrophil es are numerous in soil s close to neutrality but also occur in low numbers in acidic soil slH. It has been reported that p H optimum for hy­drolys is of co lloidal chitin is broadly related not only to the pH optimum for growth but also to the pH of the soil from wh ich the iso lates comes, which seemed in agreement to our observati ons, although some de­viati ons could be observed. The iso lates capable of max imum chitinol ys is at ac idic pH can be important fo r recycling of chitinous matter in ac idic so il s. Chiti ­nases from these isolates can be commercially ex ­ploited in biologica l control o f funga l pl ant pathogens as we ll as in fungal protoplas ting.

A lkalophilic bacteria are found w ith highest fre­quency in alka line soil s although they are also present in neutral and acidic so il s but in limited numbers. Most enzy mes from alkalophiles are characteri zed by an optimum pH on the alkaline side. A good number of iso lates capable of chitin degradat ion in the alka­line range could be iso lated in thi s study. M ost of these were bac illary but few actinomycetes al so ex ­pressed ac ti vity and grew optimall y at pH 9.0. These iso lates can be eco log ica ll y important as being most responsible members o f chitin degradation in habitats with alkaline pH and al so can be industriall y impor­tant. Chitinases w ith alkaline pH optima and stab ility can be appli cat ive in biological controI19

.2 1

. The peri­trophic membrane o f insect gut lining is chitinous and the alkali active chitinases can be used in synergism with other biocontro l agents. Chitinases acti ve and stab le at alka line pH are expected to have a better hal f-life in the gut of the insects, which has an alka­line pH, in compari son to chitin:1ses fro m neutrophi les or acidophil es, which may ex hibit a shorter half-li fe at th at pH.

The possib le role of chitin degraders in alka line habitats li ke Lonar Lake is questi onable from the poi nt of view of the source of thi s po ly mer in thi s en­vi ronment. Sal ine lakes are genera ll y dominated by blue-green algae and are poor in plankton ic and ben­thic fauna . T hi s is particularl y true for Lonar Lake, where Spirlllilla platellsis is found explaining certain leve l of eutrophicati on in the lake. Haloa lkalophili c archea can sometimes be a part of hypersa line and alkaline lakes. These lakes generally function ,IS se l f­sustain ing ecosystems due to the addition o f nutrien ts and bioe lements from creek inflow and cycling of most m;lterial internall yH There also is a poss ibility of existence of the brine shrimp like Ar/clIlia sp., fly

larvae, few c1adoceran and copepod cru staceans in these habitats.

Most efficient chitin degraders were from Mula and Mutha ri vers, agri cultural land, Jeddah and M ecca. The so il and water samples from pol luted lo­cations and garden and agri cultural soil s had more number of efficient chitinolytic bacteria. Thus, soil s and water bodies with a high organ ic content and thus high competition seem to favor the predominance of isolates expressing good chitinolytic activity. This fact may also be app li cab le to other characteristi cs of these microorganisms.

The identification of most important chitin degrad­i ng bacteria revealed streptomycetes to be the major contributors to chitin degradation irrespecti ve of ex­treme or moderate habitat. They are also predom inant in the culti vable regions further suggesting their im­portance in the bioconvers ion of chitin i!l these hab i­tats. The ability o f ACl illoplancs to survive and grow in a wide pH range explains their occurrence in alka­line regions of laundry. Further, their ability to de­grade chitin in a w ide pH range explains their possible role in chitin degradati on in alka line habitats. How­ever, potent chitinolyt ic streptomycetes were not found in Sassoon Docks and Pashan Lake. Their rel::t­tive in frequent occurrence in Sassoon Docks could be as this habitat is of marine origin . Con tradictory to Lonar Lake, Pashan Lake did not harbor potent chiti­noly tic streptomycetes despite being polluted, a fact that needs further explorat ion. The non-streptomycete actinomycetes were mainl y found in moderate loca­ti ons and their contribution to chitin degradati on in these hab itats needs further studies.

Thus from th is study it could be ascertained that chitin-degrading bacteri a may be ubiquitous relative to the abundance of thi s polymer in nature. Chitinoly­sis can occur in a wide pH range and pH as an envi­ronmental factor seems to have an impact on the dis­tribution and activity o f chitinoly tic bacteri a, up [ 0 a certain ex tent. Chitinases of bacteria from extreme locations might ex hibit some unique properties with respect to activ ity, stabilit y or other characters, wh ich might be industriall y important. Pollution may ill gen­eral affect surviva l or enrichment of chitinoly ti c bac­teri a depending 011 the nutrient ava ilab ility, a fact that st<L!lds true fo r other enzymes as well.

Acknowledgements Fi nancia l support Crom UGC, New Del hi is highly

acknow ledgeJ.

254 IND IAN J F.XP BIOL, MARCH 2003

References I Kehya ni N 0 & Roseman S, lJiocililll Biopllvs AC/o, 1473

( 1999) 108. 2 Pace N R, Sciellce , 276 ( 1997) 734. 3 Clese ri L S, Greenberg A E & Trussell R R, Alllericall Pllblic

Ncol/II Associ(lfiOIl (A PNA ) (Ameri can Water Works Asso­ciat ion and Water Pollution Control Federation. Eel. 17th APHA, Washington DC) 1989.

4 Hsu S C & Lockwood J L, Appl Microbial, 29 ( 1975) 422. 5 Williams S T & Robinson C S, J Cell Microbiol, 127 ( 19R 1) 55 . 6 Monreal J & Rcese E T, Call J Microbial . IS ( 1969) 689. 7 Willi ams S T, Sharpe M E & Holt J G, Berge.I"s MOllllal of

Svs/cl/ /(/ /ic BOCieriology. (Vol. 4. Willi ams and Wilkins, London) 1989.

B Shirling E B & Gottli eb D. III/ J Sys/ BaC/eriol. 16 ( 1966) 311.

9 Stancck. J A & Robcrts G D, Apl)1 Microbiol, 28 ( 1974) 226. 10 Langham C D, Williams S T, Sneath P H A & Mortimer A

M , J Cell MicruiJiol, 135 ( 1989) 121. II Holt J G. Kri eg N R. Sncath P H A, Staley J T & Williams

S T, Bergev 's MClllllol of De/eo llillo/ive BOCieriolagv (N inth Ed ition, Williams and Wilkins, London) 1994.

12 Nawan i N N, Kapadni s B P, Da~; A D, Rao A S & Mahaj;1I1 S K, J Appl Microbilll, 93 (2002) 965 .

13 Hara S, Yamamura Y, Fuji i Y, Mega T & Ike naKa T. J /Jill­cll elll , 105 ( 1989) 4R4.

14 Nawa ni N N & Kapadni s B p, J Appl Microliiol, 90 (200 I) 803 .

IS Broadway R M, Williams D L, Ka in C W. Ha rman G E. Lorito M & Labeda D P, Lell API'I Microbiol, 20 ( 1995) 27 1.

16 Tsujibo 1-1 , Yoshida Y, Miyamota K, J-bsegawa T & Inamori Y, /Jiosci BiOleclll101 Biocllelll. 56 ( 1992) 1304.

17 Bhushan B. J Appl Microbiol, 88 (2000) 800. 18 Williams S T & Mayficld C I, Soil Bioi Bioc/wlII. 3 ( 197 1)

197. 19 Wiwat C, Lertca nawaniclraku l M, Siwayapram P, Pantu wa­

tana S & Bhumirata na A, Celie 179 ( 1996) 11 9. 20 Sampson M N & Gooclay G W. Microbiology. 144 ( 1998)

2 189. 2 1 Bhushan B & Hoondal G S, Wo..td J Microbiol /Jio/ec/lIlOl.

IS ( 1999) 403. 22 Nandy N & Deo V, Tata Iron and Steel Company, hmshecl­

pur, 8 ( 1961) 1.