2. DISORDERS IN PROTEIN NUTRITION

2.1 Dietary essential amino acid deficiency

Although all fish examined to date display reduced growth when fed essential amino acid (EAA) deficient diets, Table 1 shows the additional gross anatomical deficiency signs which have been reported under experimental conditions with juvenile fish fed rations deficient in one or more particular EAA's.

Table 1. Reported essential amino acid (EAA) deficiency signs in fish
Limiting EAA	Fish species	Deficiency signs¹
Lysine	Oncorhynchus mykiss	Dorsal/caudal fin erosion (1,2), increased mortality (2)
	Cyprinus carpio	Increased mortality (3)
Methionine	O. mykiss	Cataract (4,5,14)
	Salmo salar	Cataract (6)
Tryptophan	O. mykiss	Scoliosis² (7–10), lordosis ² (7,10), renal calcinosis (8), cataract (7,9), caudal fin erosion, decreased carcass lipid content (9); elevated Ca, Mag, Na and K carcass concentration (7)
		Scoliosis (11)
	Oncorhynchus nerka	Scoliosis (12,13), cataract (13)³
	Oncorhynchus keta Oncorhynchus kisutch	Scoliosis (12)
Miscellaneous	C. carpio	Increased mortality and incidence of lordosis observed with dietary deficiencies of leucine, isoleucine, lysine, arginine and histidine (3)

¹ 1-Walton, Cowey & Adron (1984),
2-Ketola (1983),
3-Mazid et al. (1978),
4-Walton,Cowey & Adron (1982),
5-Poston et al. (1977),
6-Barash, Poston &Rumsey (1982),
7-Walton et al. (1984),
8-Kloppel & Post (1975),
9-Poston &Rumsey (1983),
10-Shanks, Gahimer & Halver (1962),
11-Halver & Shanks (1960),
12-Akiyama et al. (1985),
13-Akiyama, Mori & Murai (1986),
14-Cowey et el.(1992)
²Curvature of the vertebral column
³ Reported incidence of scoliosis and cataract increased with decreasing andincreasing water temperature, respectively (13)

Under practical farming conditions dietary EAA deficiencies may arise from various routes, including:

Poor feed formulation due to the use of disproportionate amounts of feed proteins with natural specific EAA deficiencies. Table 2 presents the chemical score and limiting EAAs of some selected food proteins available to the fish feed compounder. For the sake of comparison chemical scores have been calculated for individual protein sources with reference to the mean dietary EAA requirements of rainbow trout (O. mykiss) and common carp (C. carpio) as given by Ogino (1980). Compared to fish meal, which has a well balanced EAA profile, the majority of protein sources presented have amino acid imbalances which render them unsuitable as a sole source of dietary protein for fish. For example, the deficiency of methionine in plant proteins, yeast, meat and bone meal, blood meal, and hydrolysed feather meal; the deficiency of lysine in oilseeds, hydrolysed feather meal and algae; the deficiency of threonine in some oilseeds and pulses; and the deficiency of tryptophan in fish silage. It is clear from the above that during feed formulation special care must be given to the choice of feedstuffs used so that the desired overall dietary EAA profile is obtained.

Dietary imbalances may also arise from the presence of disproportionate levels of specific amino acids; including leucine/isoleucine antagonisms, and to a lesser extent arginine/lysine and cystine/methionine antagonisms. For example, blood meal is a rich source of valine, leucine and histidine, but is a very poor source of methionine and isoleucine. However, in view of the antagonistic effect of excess leucine on isoleucine, animals fed high dietary levels of blood meal suffer from an isoleucine deficiency caused by an excess of dietary leucine (Taylor, Cole & Lewis, 1977). Although similar antagonisms have also been reported for cystine/methionine (use of hydrolysed feather meal; Ichhponani & Lodhi, 1976) and arginine/lysine (Harper, Benevenga & Wohlhueter, 1970) in terrestrial farm animals, they have not been reported to occur in fish fed synthetic amino acid diet combinations (Robinson, Wilson & Poe, 1981).

Dietary EAA deficiencies may arise from excessive heat treatment of feed proteins during feed manufacture. For example during fish meal manufacture excessive heat treatment has been shown to markedly reduce protein digestibility and biological value due to the destruction of amino acids by oxidation or through the formation of linkages between individual amino acids which are more resistant to digestion (McCallum & Higgs, 1989; Pike, Andorsdottir & Mundheim, 1990). The free epsilon amino groups of lysine are particularly susceptible to heat damage, forming addition compounds with non-protein molecules (reducing sugars) present in the foodstuff (Cockerell, Francis & Halliday, 1972). In addition to decreased EAA availability, toxic substances such as gizzerosine (2-amino-9-(4-imidazolyl)-7-azanonanoic acid) may also be produced from heat treated fish meals containing free histidine and histamine (Okayaki et al., 1983; Watanabe et al. 1987). Although gizzerosine is reported to cause gizzard erosion (GE) in chicks, toxicological signs of rainbow trout fed comparable heat treated whole fish meals containing high levels of histidine and histamine included decreased stomach wall thickness, pycanosis and necrosis of gastric gland cells (Watanabe et al. 1987).

Table 2. Chemical score and limiting essential amino acids of selected protein sources ¹
Feedstuff	Ref²	THR	VAL	MET	CYS	IIS	LEU	PHE	TYR	LYS	HIS	ARG	TRP	LAA³
Chick pea	1	64^*	89	63^*	104	119	110	113	86	72	100	166	129	Met
Mung bean	1	59^*	110	54^*	48^*	127	121	124	94	79	114	123	123	Cys
Cow pea	1	65^*	103	61^*	59^*	116	116	116	100	75	127	134	129	Cys
Yellow lupin	2	66^*	81	20^*	126	117	125	85	94	64*	117	192	135	Met
Lima bean	2	84	110	57^*	74	135	118	125	106	72	112	98	106	Met
Broad bean	3	77	103	30^*	41^*	115	118	98	118	77	98	160	118	Met
Haricot bean	1	80	103	43^*	67^*	120	121	118	83	92	127	104	129	Met
Safflower	2	68^*	125	63^*	141	111	99	101	100	43*	121	181	118	Lys
Crambe	2	98	121	67^*	218	117	104	83	86	66*	104	111	200	Lys
Palm kernel	2	62^*	113	94	133	95	89	72	78	41*	98	225	311	Lys
Cottonseed	2	65^*	102	52*	118	92	94	122	89	52*	117	205	141	Lys
Sunflower	2	65^*	124	83	137	115	104	109	91	42*	119	159	165	Lys
Linseed	2	71	122	93	156	111	90	105	92	43^*	100	174	182	Lys
Sesame	2	58^*	98	109	148	91	105	86	114	33^*	114	211	153	Lys
Coconut	4	65^*	114	61^*	96	115	112	95	92	37*	81	217	123	Lys
Groundnut	4	55^*	99	39^*	133	117	100	107	117	53*	100	196	141	Met
Rapeseed	4	93	118	83	70	113	116	94	77	74	131	112	159	Cys
Soybean	4	74	101	46^*	130	128	115	105	97	76	106	123	176	Met
Potato protein meal	5	89	125	63^*	96	128	120	112	149	74	73	73	118	Met
Leaf protein meal	6	84	127	57^*	56^*	112	120	122	129	71	90	96	141	Cys
Spirulina maxima	2	87	136	2^*	30^*	159	118	105	123	55*	75	111	165	Cys
Bakers dried yeast	4	93	116	63^*	85	139	112	91	108	86	106	89	141	Met
Torulopsis utilis	4	94	118	54^*	81	144	98	137	117	84	104	86	118	Met
Bacterial SCP	7	97	134	89	59^*	115	107	115	138	71	83	84	118	Cys
Whole hen's egg	8	77	125	100	130	132	109	97	98	78	92	96	135	Thr
Fish muscle	9	83	98	98	85	108	110	80	117	101	121	97	135	Phe
Fish meal (herring)	4	76	127	109	78	117	107	80	95	89	96	111	123	Thr
Fish meal (white)	4	81	106	104	93	121	109	81	94	90	94	116	129	Thr
Fish protein concentr.	2	83	110	118	63^*	127	109	85	103	92	90	95	153	Cys
Fish silage	10	98	122	72	72	101	129	120	94	98	121	108	59*	Trp
Whole shrimp meal	2	83	97	109	85	112	106	95	105	86	73	134	106	His
Meat and bone meal	4	77	128	59^*	89	109	113	88	60*	86	100	150	88	Met
Blood meal	4	69*	158	33^*	52^*	24*	162	124	69*	89	214	62*	123	IIs
Liver meal	2	76	135	72	89	105	121	109	106	71	98	105	153	Lys
Poultry byproduct meal	4	76	125	81	141	132	123	80	60^*	71	87	134	112	Tyr
Hydrolyzed feathers	4	91	164	24^*	289	131	124	78	86	33^*	50*	147	76	Met
Earthworm meal	11	107	99	106	52^*	112	124	84	108	79	125	98	82	Cys
House fly larvae	12	75	103	72	52^*	96	90	128	218	77	127	82	147	Cys

¹ Scores based on comparison with the mean essential amino acid requirements of rainbow trout and common carp (Ogino, 1980). Mean EAA requirement (expressed as% of total EAA) being: threonine 10.6, valine 9.5, methionine 5.4, cystine 2.7, isoleucine 7.5, leucine 13.5, phenylalanine 9.5, tyrosine 6.5, lysine 16.8, histidine 4.8,arginine 11.6 and tryptophan 1.7
² Source: 1-Kay (1979), 2-Gohl (1980), 3-Bolton & Blair (1977), 4-National Research Council (1983), 5-Tunnel Avebe Starches Ltd (UK), 6-Cowey et al. (1971), 7-Methanobacter methylotrophus Unpublished data, 8-Cowey & Sargent (1972), 9-Connell & Howgate (1959), 10-Jackson, Kerr & Cowey (1984), 11-Tacon, Stafford& Edwards (1983), 12-Spinelli (1980)
* Limiting essential amino acids (present below 30% mean fish requirement)

Dietary EAA deficiencies may arise from the chemical treatment of feed proteins with acids (silage production) or alkalies, due to the loss of free tryptophan and lysine/cystine respectively (Kies, 1981).
Dietary EAA deficiencies may arise from the leaching of free and protein bound amino acids into the water. For example, Grabner, Wieser & Lackner (1981), reported the loss, through leaching, of almost all the free and about one-third of the free plus protein bound amino acids from frozen or freeze-dried zooplankton (Artemia Salina and Moina spp.) after a ten minute immersion period at 9°C. A similar rapid leakage of free amino acids (FAA) has also been observed from frozen Daphnia upon thawing; 23.1% leakage of all FAA during five minutes in water after thawing (Holm & Walther, 1988).

2.2 Toxic amino acids

Nutritional pathologies may arise from the consumption of feed proteins containing toxic amino acids or their derivatives. Feed proteins containing toxic amino acids which have been reported to have a negative effect on fish growth and feed efficiency (including eventual fish death) include the plant legumes Leucaena leucocephala (toxic nonprotein amino acid - mimosine; Jackson, Capper & Matty, 1982; Wee & Wang, 1987) and Sesbania grandiflora and Canavalia ensiformis (toxic amino acid - L-canavanine; Martinez-Palacios et al. 1988, Olvera et al. 1988).

In addition to the non-essential amino acids, certain EAA (ie. leucine) have also been reported to exert a toxic effect in fish when present in dietary excesses (Hughes, Rumsey & Nesheim, 1984; Robinson, Poe & Wilson, 1984). For example, the reported toxicity signs for a dietary excess of leucine (13.4% of diet) in rainbow trout (O. mykiss) included scoliosis, deformed opercula, scale deformities, scale loss, and spongiosis of epidermal cells (Choo et al. 1991).