1. Five multiple metal resistant ciliates viz., Stylonychia mytilus, Euplotes mutabilis, Paramecium caudatum, Oxytricha fallax and Vorticella microstoma were isolated from industrial wastewater from Kasur tanneries. The protozoans were checked for their resistance to various heavy metals viz., chromium, cadmium, copper, lead, mercury, zinc and nickel.
2. Vorticella could tolerate Pb2+ (150 ug/ml), Cu2+ (22 ug/ml), Hg2+ (16 uglm1), d2+ (22 ug/ml), Zn2+ (25 ug/ml), Ni+2 (17 ug/ml) and Cr6+ (100 ug/ml). Palramecium on the other. hand, showed resistance against Pb2+, Cu2+, Cr6+, Ni2+, Zn2+, Hg2+ and Cd2+ at a concentration of 25, I5, 20, 18, 15, 17 and 14 utg/ml, respectively. Eluplotes tolerated Pb2+ Cu2+, Cr6+ Zn+, Hg2+, Cd2+ and Ni2+ at a concentration of 75, 22, 60, 33, 22, 22 and 18 ug/ml, respectively. Stylonychia mytilus showed maximum resistance against Pb2+ i.e. 60 ug/ml and minimum resistance against Hg2+ and Ni2+(16 ug/ml). The ciliate could also resist Cu2+, C.r6+, Cd2+ and Zn2+ at a concentration of 20, 30, 23 and 30ug/ml, respectively. Oxytricha showed resistance against Pb2+, Hg2+, Ni2+, Cu2+, Cd2+, Zn2+ and Cr2+ at a concentration of 60, 18, 18, 16, 16, 17 and 50 ug/ml.
3. All ciliates studied here showed maximum growth in Bold-basal salt medium.
4. All ciliates showed optimum growth at 25°C, and pH 7.5 except Paramecium, which showed optimum growth at 30oC and Oxytricha, which showed best growth at pH 8.0.
5 While studying the seasonal population variation, it was observed that (a) Amoeba, Paramecium, Euplotes and Childonella disappeared in the industrial effluents during August (34°C) though were present in fairly large number during other months of the year. Cyclidium and Spatchidium, on the other hand, disappeared during winter month of January (17°C)
(b) Oxytricha, Amoeba, Plagiopyla and Euplotes were not observed during September when wastewater had pH 9.7. Likewise,paramecium Cyclidium and Spatchidium were not found in industrial wastewater with pH 7.71 but were found in large number at fairly high pH range (8.05-9.44)
(c) The industrial wastewater had low concentration of chromium(0.05 ug/md during March and April, which increased to1.6 ug/ml in the month of July. Euplotes.Cyelidium, plagiopyla, Tetrahymena and Euglena. Which were abundantly present in chromium concentration of 0.05 ug/ml, disappeared at 1.6 ug/ml.
6. Protozoans isolated from industrial wastewater have shown remarkable ability to pick up heavy metal ions from the culture medium. Vorticella reduced 95% of Cr+6, 98% of Pb2+, 92% of Zn2+, 94% of Cd2+, and 91 % of Hg2+ from the medium after 96 hours of incubation in a culture medium each containingl0 ug/ml of the reactive metal ions. Besides this, 94% of Cu2+ (5 ug/ml) and 86% Ni2+ (5 ug/ml) were also removed by the ciliate from the medium after 96 hours, respectively. Stylonychia could eliminate 86% of lead, 80% of chromium, 98% of zinc, 90% of mercury, 88% of Cu2+, 91 % of Cd2+ and 73% ofNi2+ from the medium after 96 hours of incubation.
Euplotes reduced 97% of lead, 98% of chromium, 84% of zinc, 82% of mercury from the medium after 96 hours and 95% of Cu21, 84% of Cd21 and 87% ofNi2+ from the medium after 6 days. Paramecium could reduce 90%, 92%, 95%, 78%, 94%, 82% and 76% of cr2+. Pb2+, Zn2+, Hg2+, Cu2+, Cd2+ and Ni2+ from the medium after 96 hours, respectively.
Oxytricha on the other hand, could remove 97%, 90%, 91 %, 90%, 94%, 88%, and 94%Cr6+, pb2+, Hg2+, Cu2+, Ni2+ , Cu2+and Cd2+ - from the medium after 96 ours, respectively.
7. PAGE analysis of total proteins of the protozoan isolates showed that (i) A number of new proteins were induced against Pb2+ (70kDa, 60kDa, 50kDa, 45kDa, I40kDa, 38kDa, 23kDa, 20kDa and 18kDa), Cr6+ (60kDa, 50kDa, 45kDa, 40kDa, 20kDa, 18kDa and 14kDa), and Cd2+ (50kDa, 45kDa, 40kDa, 30kDa, 18kDa and 9kDa). (ii) Certain proteins disappeared as a result of metal stress. For example a 55kDa protein was present in control, Pb2+ -treated and Cr6+ -treated samples but was absent in Cd2+ treated sample. Likewise another 44kDa protein was present in control, Pb2+.treated and Cr6+-treated samples but was absent in Cd2+ -treated sample.
It was concluded that multi metal-resistant ciliates had remarkable ability to pick up metal ions ions the culture medium and hence are the best candidates for removal of the various heavy metals from industrial wastewater.