Fish disease controlling efficacy study of selected …Ashokkumar R et al 178 J Pharm Chem Biol Sci, June-August 2016; 4(2): 176-187 Collection of fingerlings Fingerlings of Channa

176

J Pharm Chem Biol Sci , June-August 2016; 4(2): 176-187

Journal of Pharmaceutical, Chemical and Biological Sciences

ISSN: 2348-7658 Impact Factor (GIF): 0.615

Impact Factor (SJIF): 2.092 June-August 2016; 4(2):176-187

Fish disease controlling efficacy study of selected Indian

medicinal plant

Ashokkumar R*, M. Ramaswamy

Department of Zoology, Karpagam University, Coimbatore 21, India

*Corresponding Author: Ashokkumar R, Department of Zoology, Karpagam University, Coimbatore 21,

India

Received: 22 October 2015 Revised: 15 July 2016 Accepted: 17 July 2016

INTRODUCTION

Medicinal plants are of great importance for the

health of individual and the society. The

medicinal value of these plants lies in some

chemical substances that produce a definite

physiological action on the human and animal

body. The Indian and Chinese people depend

on medicinal plants as an alternate medicine

remedy against infections. Among various

kinds of cultivated organisms, many marine and

freshwater finfish and shellfish species

constitute an important industry with their

production increasing every year. Recently, due

to intensive farming practices, infectious

diseases pose a major problem in aquaculture

Original Research Article

ABSTRACT

The present study is aimed to investigate the disease controlling efficacy of the methanolic leaf

extract of Indian medicinal plant, Phyllanthus amarus, amended with manually prepared feed fed to

fingerlings of edible freshwater fish, Channa striatus(Bloch) which was exposed to two different

concentrations (10-5& 10-3) of two species of microorganisms, Pseudomonas fluorescens (bacteria)

and Penicillium chrysogenum (fungus)in different rearing tanks. Haematological parameters of

control (prefed with normal feed) and experimental (prefed with plant extract/chemical antibiotics)

fingerlings revealed that the methanolic extract of Phyllanthus amarushas has got higher controlling

efficacy of bacterial infection than fungal infection by way of increasing the lymphocyte count in the

blood to stimulate humoral immune response by way of producing immunoglobulins to fight against

harmful microorganisms.

Keyword: Phyllanthus amarus; Pseudomonas fluorescens; Penicillium chrysogenum; channa

striatus(Bloch); lymphocyte; humoral immune response; immunoglobulins.

Ashokkumar R et al 177


industry, causing heavy loss to farmers. In

order to address this problem, several studies

have been conducted on the modulation of fish

immune system in order to prevent the disease

outbreak as reviewed recently by Sakai (1999)

[1]. Disease outbreaks are increasingly being

recognized as a potential constraint on

aquaculture production and trade, and cause

massive financial loss through mortality or

reduced meat quality, resulting in reduced

profit margins, Smith et al., 2003 [2]. The

economic loss due to the diseases outbreak in

the aquaculture sector can be considerable.

For example, economic loss attributed due to

the diseases in the Asian region countries was

estimated to be at least US$ 1400 million in

1990 (ADB/NACA 1991). Antimicrobial effects

of different extracts of various medicinal plants

were studied by a number of workers,

Subhadradevi V et al., 2011; Anushia C et

al.,2009; Ramakrishnan G et al.,2011; Kaveri

Singh et al.,2010; Sathya A V et al.,2012;

Mahesh B and Satish S, 2008; Foysal M J et

al.,2011; Chakraborthy G S, 2008; Sathya A and

Ambikapathy V, 2012; Ogunjobi A A and

Ogunjobi T E, 2011. [3-12].

MATERIAL AND METHODS

Collection of plant

Leaf sample of medicinal plant species,

Phyllanthus amarus was collected from

Kalingarayan canal bank at Bhavani (Erode

District, Tamilnadu) Identification of the plant

species was done with the help of

Dr.R.Gopalan, Professor of Botany Karpagam

University (former Scientist, BSI,

Coimbatore),Coimbatore.

Preparation of leaf extract

The collected leaves were shade dried at room

temperature for 20 days. The dried leaves were

powdered in mechanical grinder. 20 grams of

leaf powder was weighed, 150ml of methanol

solvent was added and kept for 3 days. The

extract was filtered using Whatman No.1 filter

paper and the supernatant was collected. The

residue was again extracted two times (with 3

days of interval) and supernatants were

collected. The supernatants were pooled and

evaporated (at room temperature, 28± 1˚C) till

the volume was reduced to fully dried level.

Extract of the leaf powder of the Phyllanthus

amarus stored in air tight bottles for further

analysis.

Preparation of normal and extract/chemical

antibiotics-amended fish feeds.

The normal fish feed was prepared by using

known quantities of ingredients such as

Anchovy, Jawala (dry fish), soya flour, tapioca

flour, wheat flour and rice flour to maintain the

required protein level. The selected ingredients

were powdered and sieved to get fine particles

of uniform size. Then, the ingredients were

weighed according to the formulation [13] hand

kneaded by adding sufficient quantity of

distilled water and finally made into a dough.

The dough was then autoclaved in a closed

aluminum container at 105oC for about 15

minutes and then cooled. To the cooled dough,

aqua savor, mono sodium phosphate, vitamin

pre mix, mineral pre mix, vitamin C and

vegetable oil were added and this served as the

normal feed. The normal feed (Plate 13) thus

prepared, was divided into five parts. One part

was treated as control feed and the remaining 4

parts were used for preparing amended feeds

by mixing the methanolic leaf extract of P.

amarus, tetracycline and fluconazole

(2.5gm/50kg body weight concentrations).

Preparation of stock culture of microorganisms

Stock cultures of Pseudomonas fluorescens was

whereas, Penicillium chrysogenum was

grown in potato dextrose agar (PDA) nutrient

broth at 30˚C. Sub cultures of the

microorganisms were also prepared and

maintained at 4˚C, *14+.



Collection of fingerlings

Fingerlings of Channa striatus, the freshwater,

Indian murrel, vernacularly called as “Verral” or

“Snake headed fish”. A bulk sample of

fingerlings (of 3 ± 0.5 cm length and 10 ± 2 gm

Wt) of Channa striatus were obtained from

M/S. Murugan fish farm, Palayamkottai. The

bulk samples were acclimated for 10 days in

large fiber glass tanks with regular feeding with

normal fish feed [13].

Collection of blood samples for haematological

analysis

For haematological observations, blood sample

was collected by direct cardiac puncture of

fingerlings using a sterile insulin needle (1ml)

pre-rinsed with anticoagulant (heparin)

solution. 0.5ml of blood sample was obtained

by pooling the blood collected from 10

fingerlings for each experimental set up. Blood

samples were collected from control fingerlings

prefed with normal feed and microbe exposed,

and fingerlings prefed with amended feed and

microbe-exposed at the end of 1st, 7th, 14th and

21st days. The haematological parameters such

as total RBC count, haemoglobin content, total

leucocyte count (TLC) and differential leucocyte

count (DLC) were estimated in the blood of

fingerlings of Channa striatus prefed with

control/amended feeds and exposed to

microorganisms [13].

RESULTS

Haematological parameters in microbe-

exposed fingerlings of Channa striatus prefed

with control and amended feed

The data on the different haematological

parameters of control (prefed with normal

feed) and experimental (prefed with plant

extract/chemical antibiotics) fingerlings

exposed to two different concentrations of

microorganisms are presented in Tables 1 to 8.

Comparative analysis of percent changes (from

control level) in different haematological

parameters in the blood of experimental

fingerlings prefed with amended feed and

exposed to microorganisms are shown in Figs. 1

to 8.

(a) Total erythrocyte count

The total erythrocyte count in the blood of

control (prefed with normal feed and

Pseudomonas fluorescens exposed for 6 days)

fingerlings at the end of 21 days ranged from

3.30 ± 0.01 to 3.40 ± 0.01 106cells/mm3. The

total erythrocyte count of the fingerlings

registered elevations (ranging from 0% to 3%)

from the control levels (excepting that of 10-3

concentration of bacteria – exposed fingerlings

prefed with PA extract) (Table 1).



Table 1. Total erythrocyte count (106cells/mm3) in the blood of control (prefed with normal feed)

and experimental (prefed with plant extracts/chemical antibiotic) fingerlings exposed to two

different concentrations of Pseudomas fluorescences and Penicillium chrvsogenum.

Values are means of 6 observations ± S.E. percent from control level are given in the parenthesis

(b) Haemoglobin content

The haemoglobin content of control fingerlings

(prefed with normal feed) from 1st to 21st days

after microbial exposure insignificantly ranged

from 12.24 ± 0.06 to 13.90 ± 0.05 g/100ml.

Though the patterns of changes in haemoglobin

content showed variation up to 14th day, after

21st day, the haemoglobin content registered

elevations (ranging from 0% to 14%) in the

blood of fingerlings of all the experimental

conditions (Table 2)

Fingerling of Channa striatus

Concentration of

microorganism

Days after 6 days microbial exposure ‘F’ value

1st 7th 14th 21st

Pseudomonas fluorescens –exposed

Control (prefed with normal feed)

10-3 3.30 ± 0.01 3.31 ± 0.01 3.36 ± 0.01 3.34 ± 0.01 0.42 NS

10-5 3.04 ± 0.02 3.05 ± 0.02 3.05 ± 0.02 3.08 ± 0.02 0.86 NS

Prefed with PA extract

10-3 3.04 ± 0.01

(-8) NS 3.10 ± 0.01

(-6) NS 3.25 ± 0.01

(-3)NS 3.26 ± 0.01

(-2)NS 1.25 NS

10-5 3.08 ± 0.01 (-1) S

3.23 ± 0.01 (+6) HS

3.32±0.01 (+10)HS

3.40±0.01 (+10) HS

15.65 S

Prefed with Tetracycline

10-3 3.31 ± 0.01 (0) S

3.14 ± 0.01 (-5) NS

3.19 ± 0.01 (-5)NS

3.29 ± 0.01 (-1) NS

8.52 S

10-5 3.10 ± 0.01 (+2) S

3.18 ± 0.01 (+5) S

3.25±0.01 (+7)HS

3.40±0.01 (+12)HS

25.42 HS

Penicillium chrysogenum-exposed


10-3 3.33 ± 0.01 3.30 ± 0.01 3.35 ± 0.01 3.38 ± 0.01 0.28 NS

10-5 3.05 ± 0.02 3.07 ± 0.02 3.01 ± 0.02 3.09 ± 0.02 0.46 NS


10-3 3.11 ± 0.01

(-7) NS 3.09 ± 0.01

(-6) NS 3.20 ± 0.01

(-4)NS 3.35 ± 0.01

(-1) NS 6.48 S

10-5 3.19 ± 0.01 (+5) HS

3.00 ± 0.01 (-2) NS

3.20±0.01 (+6)HS

3.38 ± 0.01 (+9) HS

12.56 S

Prefed with Fluconazole

10-3 2.91 ± 0.0(-13) HS

2.80 ±0.01 (-15) HS

2.98 ±0.01 (-11)HS

3.30 ± 0.01 (-2) HS

3.52 S

10-5 3.20 ± 0.01 (+5) HS

3.33 ± 0.01 (+8) HS

3.32±0.01 (+10)HS

3.43±0.01 (+11)HS

14.86 S



Table 2. Haemoglobin (g/100ml) count in the blood of control (prefed with normal feed) and

experimental (prefed with plant extracts/chemical antibiotic) fingerlings exposed to two different

concentrations of Pseudomonas fluorescens and Penicillium chrysogenum.

Values are means of 6 observations ± S.E. Percent changes from control levels are given in the

parenthesis.

(c) Total leucocytes count (TLC)

The total leucocytes count of control fingerlings

of Channa striatus up to 21 days ranged

between 12.00 ± 0.04 to 12.40 ±

0.05103cells/mm3 which were found to be

statistically insignificant. However, following 21

days, fingerlings (prefed with extracts and

chemical antibiotics and microbe exposed)

registered significant elevations (over control

level) in the total leucocytes count ranging from

0% to 3% excepting that of fingerlings prefed

with fluconazole and fungus–exposed, which

showed -1% reduction from control level

(Table 3).

.


Concentration of

microorganism


1st 7th 14th 21st



10-3 12.24 ± 0.06 12.23 ± 0.06 12.25 ± 0.06 12.24 ± 0.06 0.36 NS

10-5 12.30 ± 0.05 12.42 ± 0.05 12.38 ± 0.05 12.32 ± 0.05 0.14 NS


10-3 12.12 ± 0.06(-1) NS

12.21 ± 0.05 (0) NS

12.25 ± 0.05 (0)NS

12.35 ± 0.06(+1) S

4.85 S

10-5 13.55 ±0.05(+10) HS

13.40 ± 0.06(+8)HS

13.65 ± 0.05(+10)HS

13.89 ±0.04(+13) HS

16.25 HS


10-3 12.16 ± 0.05 (-1) NS

12.12 ± 0.05(-1) NS

12.25 ± 0.04 (0) NS

12.40 ± 0.03 (+1) S

30.46 HS

10-5 13.45 ± 0.03 (-9) HS

13.65±0.03(-10) HS

14.02 ± 0.3 (+13) HS

14.10 ± 0.03(+14)HS

25.16 HS



10-3 12.20 ± 0.06 12.26 ± 0.06 12.28 ± 0.06 12.25 ± 0.06 0.74 NS

10-5 12.42 ± 0.05 12.38 ± 0.05 12.34 ± 0.05 12.36 ± 0.05 0.28 NS


10-3 12.20 ± 0.05 (0) NS

12.06 ± 0.05(-2) NS

12.20 ± 0.02 (-1) NS

12.29 ± 0.06 (0) S

42.15 HS

10-5 12.80 ± 0.05 (+3) HS

12.92 ± 0.05(+4)HS

13.69 ± 0.05(+11)HS

13.90 ± 0.05(+12)HS

36.14 HS


10-3 12.18 ± 0.06 (0) NS

12.15 ± 0.08(-1)NS

13.05 ± 0.05 (+6) HS

13.00 ± 0.05 (+6)HS

12.86 HS

10-5 12.89 ±0.08

(+4) HS 13.30 ±

0.05(+7)HS 13.60 ±

0.08(+10)HS 13.95 ±

0.02(+13)HS 25.42 HS



Table 3. Total leucocyte count (103cells/mm3) in the blood of control (prefed with normal feed)

and experimental (prefed with plant extracts/chemical antibiotic) fingerlings exposed to two

different concentrations of Pseudomonas fluorescens and Penicillium chrysogenum.


parenthesis

(d) Differential leucocyte count (DLC)

Basophil count

The basophil count in the blood of control

fingerlings insignificantly ranged from 3.00 ±

0.04 percent to 2.00 ± 0.02 percent following 1st

to 21st days. Unlike previously mentioned

parameters, the basophil count uniformly

showed significant reduction (from control

level) ranging from 0% to -67% up to 21 days

under various experimental conditions. This

reduction in basophil content, inspite of

elevated TLC level (Table 4) is noteworthy for

discussion.


Concentration of

microorganism


1st 7th 14th 21st



10-3 12.00 ± 0.04 12.01 ± 0.04 12.02 ± 0.03 12.00 ± 0.05 0.14 HS

10-5 12.30 ± 0.03 12.34 ± 0.04 12.30 ± 0.03 12.35 ± 0.04 0.16 NS


10-3 11.70 ± 0.06 (-3) NS

11.50 ± 0.07 (-4) NS

11.80 ± 0.03 (-2) NS

12.30 ± 0.05 (0) S

12.85 HS

10-5 11.20 ± 0.02 (-9) NS

11.75 ± 0.02 (-5) NS

12.20 ± 0.05 (-1) NS

12.40 ± 0.05 (0) S

16.46 S


10-3 11.00 ± 0.08 (-8) NS

11.10 ± 0.07 (-8) NS

11.50 ± 0.07 (-4) NS

12.00 ± 0.05 (0) S

12.48 S

10-5 11.10 ±0.05(-10) NS

11.20± 0.05 (-9) NS

11.80 ± 0.02 (-5) NS

12.40 ± 0.02 (0) S

22.45 HS



10-3 12.08 ± 0.02 12.07 ± 0.03 12.11 ± 0.03 12.09 ± 0.02 0.19 NS

10-5 12.28 ± 0.04 12.32 ± 0.06 12.35 ± 0.02 12.40 ± 0.03 0.15 NS


10-3 11.40 ± 0.06 (-6) NS

11.30 ± 0.05 (-6) NS

11.90 ± 0.05 (-2) NS

12.20 ± 0.01 (+1) S

14.25 S

10-5 11.50 ± 0.05 (-6) NS

11.10 ±0.01 (-10) NS

11.70 ± 0.01 (-5) NS

12.30 ± 0.02 (+1) NS

6.45 S


10-3 11.20 ± 0.01 (-7) NS

11.50 ± 0.05 (-5) NS

11.91 ± 0.03 (-2) NS

12.50 ± 0.03 (+3) HS

24.52 HS

10-5 11.50 ± 0.02 (-6) NS

11.90± 0.06 (-3) NS

12.11 ± 0.03 (-2) NS

12.20 ± 0.05 (-2) NS

8.52 S



Table 4. Basophil count (percentage) in the blood of control (prefed with normal feed) and




parenthesis.

Neutrophil content

The neutrophil count of control fingerlings

insignificantly ranged from 40.02 ± 1.12 percent

to 33.00 ± 1.12 percent following 21 days of

experimental period. On the other hand,

fingerlings prefed with leaf extract/fluconazole

and exposed to Penicillium chrysogenum

showed significant reduction (from control level

ranging from -3% to -18%) in the neutrophil

content after 21 days (Table 5).


Concentration of

microorganism


1st 7th 14th 21st



10-3 3.00 ± 0.04 3.06 ± 0.04 3.04 ± 0.04 3.05 ± 0.04 0.46 NS

10-5 3.12 ± 0.01 3.08 ± 0.01 3.02 ± 0.01 3.03 ± 0.01 0.28 NS


10-3 3.00 ± 0.01 (0) NS

2.00 ± 0.01 (-35) NS

3.00 ± 0.01 (-1) NS

3.00± 0.02 (-2) NS

3.62 S

10-5 3.00 ± 0.01 (-4) NS

2.00 ± 0.01 (-35) NS

3.00 ± 0.01 (-1) NS

3.00 ± 0.01 (-1) NS

4.86 S


10-3 2.00 ± 0.01 (-33)NS

2.00 ± 0.02 (-35) NS

1.00 ± 0.01 (-67) NS

2.00 ± 0.01 (-34) NS

12.58 HS

10-5 3.00 ± 0.01 (-4) NS

4.00 ± 0.01 (-30) HS

2.00 ± 0.01 (-34) NS

3.00 ± 0.01 (-1) NS

3.64 S



10-3 3.00 ± 0.04 3.08 ± 0.04 3.02 ± 0.04 3.04 ± 0.04 0.52 NS

10-5 3.10 ± 0.01 3.09 ± 0.01 3.04 ± 0.01 3.06 ± 0.01 0.65 NS


10-3 3.00 ± 0.01

(0) NS 2.00 ± 0.01

(-35) NS 2.00 ± 0.01

(-34) NS 2.00 ± 0.01

(-34) NS 42.64 HS

10-5 3.00 ± 0.01 (-3) NS

3.00 ± 0.01 (-3) NS

2.00 ± 0.02 (-34) NS

2.00 ± 0.01 (-35) NS

36.48 HS


10-3 2.00 ± 0.03

(-33)NS 2.00 ± 0.03

(-35) NS 2.00 ± 0.05

(-34) NS 1.00 ± 0.02

(-67) NS 48.42 HS

10-5 2.00 ± 0.04 (-35)NS

2.00 ± 0.03 (-35) NS

2.00 ± 0.03 (-34) NS

2.00 ± 0.06 (-35) NS

26.36 HS



Table 5. Neutrophil (percentage) count in the blood of control (prefed with normal feed) and




parenthesis.

Eosinophil count

The eosinophil count of control fingerlings

insignificantly ranged from 2.00 ± 0.06 percent

to 1.00 ± 0.01 percent following 21 days of

experimental period. Unlike the neutrophil

count, both under Pseudomonas fluorescens

and Penicillium chrysogenum exposures, the

experimental fingerlings uniformly showed -

50% reductions (from control level) following

21 days under all the experimental conditions

(Table 6).


Concentration of

microorganism

Days after 6 days microbial exposure ‘F’ value 1st 7th 14th 21st



10-3 40.02 ± 1.12 40.06 ± 1.12 40.04 ± 1.12 40.05 ± 1.12 0.16 NS

10-5 40.04 ± 2.47 40.10 ± 2.47 40.08 ± 2.47 40.06 ± 2.47 0.25 NS


10-3 44.00 ± 1.32 (+10)S

39.00 ± 1.14 (-3) NS

43.00 ± 1.11 (+7)S

46.00 ± 1.20 (+15) S

4.28 S

10-5 49.00 ± 1.55 (+22)S

48.00 ± 2.05 (+19) S

49.00 ± 1.12 (+22) S

51.00± 1.11 (+27) HS

22.65 HS


10-3 44.00 ± 1.13 (+10) S

50.00 ± 1.43 (+25) HS

59.00 ± 1.32 (+47) HS

54.00 ± 1.33 (+35) HS

35.48 HS

10-5 55.00 ± 1.66 (+37) HS

52.00 ±1.21 (+30) HS

53.00 ± 1.22 (+32) HS

48.00± 2.02 (+20) S

22.87 HS



10-3 40.05 ± 1.12 40.07 ± 1.12 40.08 ± 1.12 40.11 ± 1.12 12.46 S

10-5 40.02 ± 2.47 40.04 ± 2.47 40.08 ± 2.47 40.06 ± 2.47 8.52 S


10-3 36.00 ± 2.00

(-10) NS 44.00 ± 2.01

(+9)S 37.00± 1.13

(-8) NS 33.00 ±1.53

(-18) NS 12.56 S

10-5 39.00 ± 1.45 (-3) NS

34.00 ± 2.00 (-15) NS

35.00 ± 1.99 (-13) NS

33.00 ± 1.12 (-18) NS

14.62 S


10-3 39.00 ± 2.04 (-3) NS

36.00 ± 1.33 (-10) NS

39.00 ± 1.32 (-3) NS

41.00 ± 1.41 (+2) S

6.65 S

10-5 40.00 ± 1.11 (0) NS

39.00 ± 1.20 (-3) NS

38.00 ± 1.08 (-5) NS

37.00 ± 1.12 (-8) NS

7.28 S



Table 6. Eosinophil (percentage) count in the blood of control (prefed with normal feed) and




parenthesis.

Monocyte count

The monocyte count in the blood of control

fingerlings of Channa striatus after 21 days of

experimental period following microbial

exposure insignificantly ranged from 4.02 ± 0.02

percent to 2.00 ± 0.02 percent (Table 7). From

Table 7, it could be observed that the monocyte

count in the blood of experimental fingerlings

showed reductions (of different magnitudes)

following 1st, 7th, 14th and 21st days after

microbial exposure. The reduction percentages

of different experimental fingerlings

significantly ranged from -25% to -51% (Table

7).


Concentration of

microorganism


1st 7th 14th 21st



10-3 2.00 ± 0.06 2.04 ± 0.06 2.05 ± 0.06 2.03 ± 0.06 0.46 NS

10-5 2.08 ± 0.03 2.05 ± 0.03 2.09 ± 0.03 2.08 ± 0.03 0.35 NS


10-3 1.00 ± 0.01 (-50) NS

2.00 ± 0.01 (-2) NS

1.00 ± 0.01 (-51) NS

1.00 ± 0.01 (-51)NS

28.42 HS

10-5 1.00 ± 0.01

(-52) NS 2.00 ± 0.02

(-2) NS 1.00 ± 0.01

(-52) NS 1.00 ± 0.01

(-52)NS 16.52 S


10-3 2.00 ± 0.01 (0) NS

1.00 ± 0.01 (-51)NS

1.00 ± 0.02 (-52) NS

1.00 ± 0.06 (-52)NS

27.48 HS

10-5 2.00 ± 0.01 (0) NS

2.00 ± 0.01 (0) NS

1.00 ± 0.01 (-50) NS

1.00 ± 0.01 (-50)NS

18.52 HS



10-3 2.02 ± 0.06 2.12 ± 0.06 2.08 ± 0.06 2.06 ± 0.06 0.92 NS

10-5 2.04 ± 0.03 2.05 ± 0.03 2.03 ± 0.03 2.05 ± 0.03 0.85 NS


10-3 2.00 ± 0.01 (-1) NS

2.00 ± 0.01 (-6) NS

1.00 ± 0.01 (-52) NS

1.00 ± 0.01 (-51)NS

22.15 HS

10-5 1.00 ± 0.01 (-51) NS

2.00 ± 0.01 (-2) NS

1.00 ± 0.01 (-51) NS

1.00 ± 0.01 (-51)NS

39.46 HS


10-3 1.00 ± 0.01 (-50) NS

2.00 ± 0.05 (-6) NS

1.00 ± 0.01 (-52) NS

1.00 ± 0.03 (-51)NS

46.45 HS

10-5 1.00 ± 0.05

(-51) NS 1.00 ± 0.03

(-51)NS 1.00 ± 0.05

(-51) NS 1.00 ± 0.04

(-51)NS 29.74 HS



Table 7. Monocyte (percentage) count in the blood of control (prefed with normal feed) and




parenthesis.

Lymphocyte count:

The lymphocyte count in the blood of control

fingerlings up to 21 days after microbial

exposure insignificantly ranging from 51.00 ±

2.03 percent to 62.00 ± 1.42 percent.

Experimental fingerlings exposed to

Pseudomonas fluorescensuniformly showed

reductions (from control level) in the

lymphocyte count ranging from 0% to - 47%

following 1st, 7th, 14th and 21st days. On the

other hand, experimental fingerlings exposed to

Penicillium chrysogenum registered elevations

in the lymphocyte count ranging from 0% to

+22% following all the experimental periods

(Table 8).


Concentration of

microorganism


1st 7

th 14th 21

st



10-3 4.02 ± 0.02 4.06 ± 0.02 4.09 ± 0.02 4.05 ± 0.02 0.46 NS

10-5 4.03 ± 0.04 4.05 ± 0.04 4.00 ± 0.04 4.02 ± 0.04 0.92 NS


10-3 2.00 ± 0.01 (-50) NS

3.00 ± 0.01 (-26)NS

2.00 ± 0.01 (-51)NS

2.00 ± 0.01 (-51) NS

40.25 HS

10-5 2.00 ± 0.01 (-50) NS

3.00 ± 0.01 (-26)NS

2.00 ± 0.02 (-50)NS

2.00 ± 0.02 (-50) NS

32.46 HS


10-3 1.00 ± 0.01 (-75) NS

2.00 ± 0.01 (-51)NS

2.00 ± 0.02 (-50)NS

3.00 ± 0.02 (-26) NS

12.46 S

10-5 1.00 ± 0.01 (-75) NS

2.00 ± 0.01 (-51)NS

2.00 ± 0.01 (-50)NS

3.00 ± 0.02 (-25) NS

10.52 S



10-3 4.03 ± 0.02 4.16 ± 0.02 4.09 ± 0.02 4.05 ± 0.02 0.28 NS

10-5 4.02 ± 0.04 4.04 ± 0.04 4.08 ± 0.04 4.06 ± 0.04 0.42 NS


10-3 3.00 ± 0.01 (-26) NS

2.00 ± 0.01 (-52)NS

2.00 ± 0.01 (-51)NS

3.00 ± 0.01 (-26) NS

8.52 S

10-5 2.00 ± 0.01 (-50) NS

3.00 ± 0.01 (-26)NS

2.00 ± 0.01 (-51)NS

2.00 ± 0.01 (-51) NS

42.56 HS


10-3 2.00 ± 0.02 (-50) NS

2.00 ± 0.01 (-52)NS

2.00 ± 0.03 (-51)NS

2.00 ± 0.08 (-51) NS

47.45 HS

10-5 2.00 ± 0.03 (-50) NS

2.00 ± 0.03 (-50)NS

2.00 ± 0.06 (-51)NS

2.00 ± 0.04 (-51) NS

23.56 HS



Table 8. Lymphocyte (percentage) count in the blood of control (prefed with normal feed) and




parenthesis.

DISCUSSION

Among the hematological parameters,

significant increases were observed in the

population of neutrophils and lymphocytes in

the blood of fingerlings exposed to

microorganisms and fed with feed amended

with the methanol extract of the Phyllanthus

amarus medicinal plant compared to absence

of significant change in the above said

parameters in fingerlings treated with

microorganism and fed with feed amended

with chemical antibiotics. This probably

indicates the possible immunostimulant

potential of the plant extract by way of

stimulating humoral mediated immune

response towards microbial disease control in

fish. Absence of significant increase in

monocytes in fingerlings exposed to

microorganisms and fed with feed amended

with plant extract probably indicate the

absence of cell mediated immune response in

the fish.

CONCLUSION

From the results obtained, it is concluded that

the haematological parameters of control


Concentration of

microorganism

Days after 6 days microbial exposure ‘F’ value 1st 7th 14th 21st



10-3 51.00 ± 2.03 51.04 ± 2.02 51.06 ± 2.02 51.10 ± 2.06 0.28 NS

10-5 51.04 ± 2.02 51.08 ± 2.02 51.09 ± 2.02 51.03 ± 2.02 0.42 NS


10-3 50.00 ± 2.13 (-2) NS

54.00 ± 1.16 (6) S

51.00 ± 1.22 (0) NS

48.00 ± 2.18 (-6) NS

3.56 S

10-5 45.00 ± 1.15 (-12) NS

45.00 ± 1.16 (-12)NS

45.00 ± 2.00 (-12)NS

43.00 ± 1.89 (-16) NS

6.42 S


10-3 51.00 ± 1.22

(0) NS 45.00 ± 1.12

(-12)NS 37.00 ± 1.04

(-28)NS 40.00 ± 2.00

(-22) NS 25.42 HS

10-5 39.00 ± 1.12 (-24) NS

40.00 ± 1.11 (-22)NS

42.00 ± 2.00 (-18)NS

45.00 ± 2.10 (-12) NS

22.58 HS



10-3 51.02 ± 2.02 51.04 ± 2.02 51.06 ± 2.02 51.09 ± 2.02 0.45 NS

10-5 51.05 ± 2.02 51.03 ± 2.02 51.04 ± 2.02 51.01 ± 2.02 0.16 NS


10-3 56.00 ± 2.21 (+10) S

50.00 ± 2.33 (-2) NS

58.00± 2.11 (+14)S

61.00 ±1.32 (+19) S

12.62 HS

10-5 55.00 ± 1.82 (+8) S

57.00 ± 1.13 (+12) S

60.00 ± 1.32 (+18) S

62.00 ± 1.42 (+22) HS

14.52 HS


10-3 56.00 ± 2.00 (+10) S

58.00 ± 1.21 (+14) S

56.00 ± 1.12 (+10) S

55.00 ± 1.31 (+8) S

3.58 S

10-5 55.00 ± 1.10

(+8) S 56.00 ± 1.43

(+10) S 57.00 ± 2.01

(+12) S 58.00 ± 1.02

(+14) S 6.72 S



(prefed with normal feed) and experimental

(prefed with plant extract/chemical antibiotics)

fingerlings revealed the methanolic extract of

Phyllanthus amarusto to controlling efficacy of

bacterial infection than fungal infection and the

increases of lymphocyte to stimulate humoral

immune response by way of producing

immunoglobulins to fight against harmful

microorganisms.

CONFLICT OF INTEREST STATEMENT

The authors declare that they have no conflict

of interests.

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Cite this article as: Ashokkumar R, M. Ramaswamy. Fish disease controlling efficacy study of selected Indian

medicinal plant. J Pharm Chem Biol Sci 2016; 4(2):176-187.

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Fish disease controlling efficacy study of selected …Ashokkumar R et al 178 J Pharm Chem Biol Sci, June-August 2016; 4(2): 176-187 Collection of fingerlings Fingerlings of Channa