Waheeda Raana

Isolation And Characterization Of Clostridium Perfringens From Domestic Animals An Man In Punjab - 2007

The objectives of present investigation were to isolate the Cl. perfringens from the domestic and zoo animals and human beings; characterize it through biotyping and pathogencity observation, and to develop a vaccine- from the common CI. perfringens isolate. For this purpose a total of 1240 samples of morbid tissues (faecal samples from animals and gangrenous tissues from humans). From cattle (n=180), goats (n=180), horses (n=250), camel (n=250), deer (n=28), wild beast (n=07), monkeys (n16), zebra (n10), elephant (n01), yaks (n=09), foxes (n07), jackals (n=08), baboons (n=08), and bears (n08) were collected and processed for isolation of CI. perfringens. In addition a total of 100 human cases; 83 wound swabs and 17 gas gangrene were also collected and analyzed bacteriologically. This study has indicated that Clostridium (Cl.) perfringens causes multiple clinical problems in animals and human beings as was indicated by good rate of its isolation from the examined morbid tissues and fecal samples. Of the total 1240 samples from various types of animals 297 (23.95%) indicated the presence of CI. perfringens. The overall isolation percentages of various types of CI. perfringens from the cattle, sheep goat, horses, camel, wild beast, deer, bear, jackal, zebra, monkeys, yak, elephant, baboon, foxes, and humans were 22.2, 12.2, 57.2, 8.0, 21.6, 57.1, 30.76, 37.50, 50.0, 50.0, 37.50, 33.33, 100.00 75.00, 57.14 and 18.00, respectively. Of the tested population of domestic animals, goats indicated the highest Cl. perfringens (57.2%) infection rate. In the zoo animal population, the elephant, baboons, wild beast, jackals, and foxes were shown to be heavily infected with various CI. perfringens types compared to other wild life animals species. Of the 298 isolates obtained through this investigation Cl. perfringens type D was obtained from 118 (39.7%) morbid samples of the domestic and zoo animals; CI. perfringens type A from 63 (21 .21%) samples, Cl. perfringens type B from 95 (31.98%) samples; and the CI. perfringens type E was isolated from 21(7.07%) samples. None of the samples indicated the presence of CI. perfringens type C. Of the total 100 samples from the humans, CI. perfringens type A was isolated from 14 (14%) and Cl. perfringens type D was isolated from 04 (4%). None of the human samples showed the presence of Cl. perfringens types B, C, or E. Of the 17 human gangrene tissue samples, Cl. perfringens type A was isolated from 09 (52.94%) samples and the Cl. perfringens type D was recovered from 02 (11 .76%) samples. However, all attempts to isolate Cl. perfringens types B, C or E from the human gangrene tissue/material samples were unsuccessful.

The overall findings indicated that of the total 297 samples positive for various Cl. perfringens types 63 indicated the presence of Cl. perfringens type A. Of those 63 Cl. perfringens type A isolates, 49 were recovered from the animals; and 14 were isolated from the wound swabs and gangrene tissue material samples from humans. Of the 63 Cl. perfringens type A isolates from the animals, 5 were isolated from cattle; 3 from sheep, 20 from goats; 5 from the horses; 10 from camels, 01 from the deer; 01 from the zebra, 01 from baboon, 01 from fox, 01 from the monkey, and 01 isolate was recovered from yak. Of the 14 isolates of Cl. perfringens type A from humans, 05 were recovered from the open wound swabs, and 09 strains of the organism were isolated from the gangrenous tissue material.

Of the 297 samples positive for various Cl. Perfringens types, 95 animal samples indicated the presence of Cl. perfringens type B. These 95 isolates were obtained from cattle (n=22), sheep (n=10), goats (n=30), horses (n=03), camel (n=14), deer (n03), wild beast (n=02), monkey (n=02), zebra (n=02), yak (n=01), fox (n01), jackals (n02), baboon (n02) and bear (n=02). None of the human samples was positive for Cl. perfringens type B. Isolation of C/. perfringens type B from the zoo animals is a matter of concern for the human health, as the zoo visitors have the possibility to get infected with this organism.

Of the total 297 positive samples of faecal and morbid tissues from various types of animals and human being Cl. perfringens type D isolates were recovered from 118 (39.7%) samples. Of these 118 isolates of Cl. perfringens typeD, 114 were obtained from various types of animals, and 04 isolates were from the humans. Of the 114 animal isolates, 10 from the cattle, 5 from the sheep, 44 from the goats, 9 from the horses, 27 from the camel, 4 from the deer, 02 from the wild beast, 02 from the monkey, 02 from the zebra, 01 from the elephant, 01 from the yak, 02 from the fox, 02 from the jackals, 02 from the baboon, and 01 isolate the bear. A total of 04 CI. perfringens type D isolates were recovered from gangrenous tissue and open wound samples from human beings.

During this investigation 21 isolates of CI. perfringens Type E were obtained from domestic and zoo animals. Of the 21 isolates, 03 were from cattle, 04 from sheep, 09 from goats and 03 from horses, 01 from monkey, and 01 from the baboon. All the 21 isolations were from the fecal material of above mentioned animals. None of the human samples was positive for CI. perfringens type E.

Alpha toxin was produced by all of the 63 Cl. perfringens type A isolates. Within the toxin producing isolates, there was no difference in the quality of toxin in respect to its lethality for mice, dermonecrosis effects for guinea pigs and cytotoxicity in the HeLa cells. The 07 fecal isolates were hemolytic, lecithinase (+), and positive for all biochemical characteristics of Cl. perfringens. Those isolates were not lethal for mice, indicated no dermonecrotic activity in guinea pig, and produced mild degree of cytotoxicity in the cell cultures.

The activity of beta toxin obtained from 95 isolates of CI. perfringens type B isolates was determined using standard toxin-antitoxin test carried in mice and the standard serum neutralization test with antitoxin raised in rabbits. Within the toxin producing isolates, no difference was seen in the potential of toxin based on its lethality for mice.

Epsilon toxin activity of the 114 isolates of CI. perfringens type D from animals and 4 of the human isolates was also determined. Of the 114 animal isolate, 110(96.49%), and all the 4 human isolates produced E-toxin. There was no difference in the lethal potential of toxin for mice, dermonecrotis action in guinea pig and production of CPE in VERO cells.

Iota (i) toxin activity of the 21 isolates of Cl. perfringens type E was also determined serum neutralization test in mice. Many isolates produced more than one major toxin. Ci. perfringens (CP) type

A produced Alpha (a) toxin; CP type B produced Alpha (a), Beta (3) and Epsilon (E) toxins; OP type D isolates produced Alpha (a) and Epsilon (E) toxins, and OP type E isolates produced Alpha (a) toxin + Iota (i) toxin.

The immunobiologic studies of isolates showed that many of the isolates were quite antigenic. Isolates of CI. perfringèns type D and B were found highly immunogenic as those isolates producing SN titer of 1:320.



Department of Microbiology
Phd. thesis

0998,T


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