Lysozymes in the animal kingdom

Lysozymes (EC 3.2.1.17) are hydrolytic enzymes, characterized by their ability to cleave the β-(1,4)-glycosidic bond between N-acetylmuramic acid and N-acetylglucosamine in peptidoglycan, the major bacterial cell wall polymer. In the animal kingdom, three major distinct lysozyme types have been identified — the c-type (chicken or conventional type), the g-type (goose-type) and the i-type (invertebrate type) lysozyme. Examination of the phylogenetic distribution of these lysozymes reveals that c-type lysozymes are predominantly present in the phylum of the Chordata and in different classes of the Arthropoda. Moreover, g-type lysozymes (or at least their corresponding genes) are found in members of the Chordata, as well as in some bivalve mollusks belonging to the invertebrates. In general, the latter animals are known to produce i-type lysozymes. Although the homology in primary structure for representatives of these three lysozyme types is limited, their three-dimensional structures show striking similarities. Nevertheless, some variation exists in their catalytic mechanisms and the genomic organization of their genes. Regarding their biological role, the widely recognized function of lysozymes is their contribution to antibacterial defence but, additionally, some lysozymes (belonging to different types) are known to function as digestive enzymes.     

A laboratory study of host use by the cuckoo catfish <Emphasis Type="Italic">Synodontis multipunctatus</Emphasis>

The only known non-avian vertebrate obligate brood parasite is the cuckoo catfish (Synodontis multipunctatus), a Lake Tanganyikan endemic. The cuckoo catfish parasitizes Tanganyikan mouthbrooding cichlids, and under captive conditions, will also parasitize cichlids from other Rift Valley lakes. Here we examine the frequency of parasitism by the cuckoo catfish of Ctenochromis horei from Lake Tanganyika and three species from Lake Malawi and the greater Lake Victorian system in a laboratory setting. C. horei was parasitized significantly less (17%) than the allopatric species Haplochromis latifasciatus, Haplochromis nubilus, and Metriaclima estherae (combined parasitism rate of 28%). The lower rates of parasitism in C. horei may be due to differences in the mating ritual, oviposition (e.g., long periods of pseudo-spawning before actual oviposition), and behavioral adaptations (e.g., increased aggression towards the cuckoo catfish). The number of catfish eggs per parasitized brood was similar between C. horei, H. latifasciatus, H. nubilus, and M. estherae. Our results are comparable to findings from the field for C. horei parasitism frequency and number of cuckoo catfish per brood. We also analyzed the parasitism rate of the albino morph of Metriaclima zebra, a domestic strain. Parasitism rates and number of catfish per brood were the highest in the albino morphotype suggesting that the higher levels of parasitism may be related to lower aggressive behavior, lower visual acuity, or captive influence. Cuckoo catfish and mouthbrooding cichlids provide a model system for testing brood parasitism in a laboratory setting.     

Invertebrate lysozymes: Diversity and distribution, molecular mechanism and in vivo function

Lysozymes are antibacterial enzymes widely distributed among organisms. Within the animal kingdom, mainly three major lysozyme types occur. Chicken (c)-type lysozyme and goose (g)-type lysozyme are predominantly, but not exclusively, found in vertebrate animals, while the invertebrate (i)-type lysozyme is typical for invertebrate organisms, and hence its name. Since their discovery in 1975, numerous research articles report on the identification of i-type lysozymes in a variety of invertebrate phyla. This review describes the current knowledge on i-type lysozymes, outlining their distribution, molecular mechanism and in vivo function taking the representative from Venerupis philippinarum (formerly Tapes japonica) (Vp-ilys) as a model. In addition, invertebrate g-type and ch-type (chalaropsis) lysozymes, which have been described in molluscs and nematodes, respectively, are also briefly discussed.     

Molecular characterization and expression analysis of <Emphasis Type="Italic">Hsp90</Emphasis> in <Emphasis Type="Italic">Schizothorax prenanti</Emphasis>

Aquatic animals suffer from various environmental stresses because the aquatic environment is a very complex system. To monitor the health status of fish, Hsp90 a potential early warning marker was determined in Schizothorax prenanti after infection with a bacterium. In this study, we cloned Hsp90 from S. prenanti for the first time. The full-length cDNA sequence of SpHsp90 was 2663 bp, contains an open reading frame of 2181 bp, and has a gene encoding 726 amino acids, an estimated molecular mass of 83.38 kDa, and a theoretical isoelectric point of 4.91. The SpHsp90 amino acid sequence has five conserved HSP90 family signatures and shares 87.0–95.5 % identity with other vertebrates. Phylogenetic analysis and structure comparison indicated that SpHsp90 should be a β isoform of the HSP90 family. SpHsp90 was ubiquitously expressed in all examined tissues, and the highest level of expression was in the kidney. After Streptococcus agalactiae infection, the level of SpHsp90 expression had significant changes (P < 0.05) in the hepatopancreas, spleen, kidney, and blood. The expression increased to the highest level at 6 h in the blood and at 24 h in the hepatopancreas, spleen, and kidney. The results suggested that the SpHsp90 gene could be induced by S. agalactiae in S. prenanti and that SpHsp90 may be involved in resistance to bacterial infection and provide an early warning information. The kidney is the most suitable for detecting SpHsp90 after bacterial infection.     

Characterization of Antimicrobial Peptides Isolated from the Processing by-Products of African Catfish <Emphasis Type="Italic">Clarias gariepinus</Emphasis>

Antimicrobial peptides (AMPs) as components of innate immunity system have been isolated from fish and other species. In this study, the crude proteins extracted with gradient ammonium sulfate precipitation technique from the processing by-products of African catfish Clarias gariepinus (C. gariepinus) were purified by size-exclusion chromatography and all the four obtained fractions, Clarias antimicrobial peptides I(CAP-I), CAP-II, CAP-III and CAP-IV, showed antimicrobial activity. Among of these fractions, CAP-IV showed the highest antimicrobial activity against Staphylococcus aureus, Aeromonas sobria, Aeromonas hydrophila, Escherichia coli by agar diffusion plate test and the diameter of inhibition zone was 8.34, 9.27, 6.76, 6.13 mm, respectively. The molecular weight of main peptides of CAP-IV was around 4.1 KD by SDS-PAGE analysis. CAP-IV showed antimicrobial activity against both gram-negative and gram-positive bacterial pathogens at minimum inhibitory concentrations (MICs) ranging from 105 to 420 μg/mL. The antimicrobial activity of CAP-IV was stable at wide pH range, 3–11 and was also heat-stable when temperature was below 80 °C. Freeze-thawing treatment also only had slight effects on the antimicrobial activity of CAP-IV. Besides, CAP-IV was not sensitive to the hydrolysis by pepsin and trypsin, except for protease K. These results suggest that CAP-IV isolated from C. gariepinus is potential to be developed as a new antimicrobial peptide and may partially explain the high disease resistance of African catfish C. gariepinus.     

Stability,oviposition attraction,and larvicidal activity of binary toxin from <Emphasis Type="Italic">Bacillus sphaericus</Emphasis> expressed in <Emphasis Type="Italic">Escherichia coli</Emphasis>

Bacillus sphaericus produces a two-chain binary toxin composed of BinA (42 kDa) and BinB (51 kDa), which are deposited as parasporal crystals during sporulation. The toxin is highly active against Culex larvae and Aedes and Anopheles mosquitoes, which are the principal vectors for the transmission of malaria, yellow fever, encephalitis, and dengue. The use of B. sphaericus and Bacillus thuringiensis in mosquito control programs is limited by their sedimentation in still water. In this study, the binA and binB genes were cloned and the recombinant BinAB protein was expressed in three strains of Escherichia coli. These recombinant strains were used in a toxicity assay against Culex quinquefasciatus larvae. The highest expression level was achieved when both proteins were expressed in a single operon construct. The BinAB protein expressed in the E. coli Arctic strain showed higher larvicidal activity than either of the recombinant proteins from the E. coli Ril or pLysS strains. Furthermore, it had the highest oviposition attraction (49.1%, P?相似文献   

Trematodes (Plathelminthes) of <Emphasis Type="Italic">Clarias gariepinus</Emphasis> (Pisces: Clariidae) in Lake Tana,Ethiopia

All of the 166 Clarias gariepinus catfishes in Lake Tana, Ethiopia, were examined for trematode infestation in 2006—2009. Seven trematode species—Eumasenia bangweulensis, Astiotrema reniferum, Orientocreadium batrachoides, Paralecithodendrium chilostomum, Phyllodistomum bavuri, P. tana, and Cladorchiidae gen. sp.—as adult were found. The common catfish parasites were Eumasenia bangweulensis (20% prevalence and 1—62 intensity of invasion), Orientocreadium batrachoides (30% prevalence and 1–31 intensity of invasion), Phyllodistomum bavuri (24.8% prevalence and 1–8 intensity of invasion), Ph. tana (17.6% prevalence and 1–23 intensity of invasion), and Ph. bavuri. Astiotrema reniferum (three specimens were only found) was a rare species; Paralecithodendrium chilostomum was an accidental parasite of catfish. All these trematodes were first recorded in Ethiopia and Eastern Africa.     

Lysozyme activity in animal extracts after sodium dodecyl sulfate-polyacrylamide gel electrophoresis

1. Lysozyme activity was detected after electrophoresis in sodium dodecyl sulfate-polyacrylamide gels containing 0.2% (W/V) autoclaved Micrococcus lysodeikticus cells as substrate. 2. Lysozyme activity appeared as clear lysis zones after incubation of opaque gels at 37 degrees C in buffered Triton X-100. 3. As low as 0.1 pg of purified hen egg white lysozyme could be detected after 16 hr incubation at pH 6.5. 4. Bands with lytic activity from kidney and pancreas acetone powders, bird's egg whites and vitelline membranes, animal sera and human saliva corresponded to c-type (Mr 14,500), g-type (Mr 20,500) or both lysozymes as far as molecular weight is concerned. 5. Some extracts, like porcine kidney, exhibited more than two bands. 6. Bands with lytic activity migrating at the level of g-type lysozymes were detected in some kidney and pancreas extracts.     

Enzymatic depolymerization of <Emphasis Type="Italic">N</Emphasis>-succinylchitosan

A complex of chitinolytic enzymes of Streptomyces kurssanovii and also lysozyme and Celloviridin, an industrial cellulase preparation, were demonstrated to provide for an enzymatic hydrolysis of N-succinylchitosan. Our studies were carried out on a high-molecular N-succinylchitosan with M of 390 kDa and a substitution degree of 0.8 as a substrate. All the enzymatic preparations were shown to be suitable for the obtaining of low-molecular derivatives of N-succinylchitosan. The complex of enzymes from S. kurssanovii showed the greatest activity: they reduced the characteristic viscosity of initial solution of the substrate by 78% for 30 min. A biodegradation of N-succinylchitosan of various molecular masses was shown to proceed under the action of lysozyme, and the cleavage reaction was revealed to decelerate at a decrease of the polymer molecular mass. A use of N-succinylchitosan in a complex with drugs for a prolongation of their action in a live organism was presumed.     

Role of Lysozyme Inhibitors in the Virulence of Avian Pathogenic Escherichia coli

Lysozymes are key effectors of the animal innate immunity system that kill bacteria by hydrolyzing peptidoglycan, their major cell wall constituent. Recently, specific inhibitors of the three major lysozyme families occuring in the animal kingdom (c-, g- and i-type) have been discovered in Gram-negative bacteria, and it has been proposed that these may help bacteria to evade lysozyme mediated lysis during interaction with an animal host. Escherichia coli produces two inhibitors that are specific for c-type lysozyme (Ivy, Inhibitor of vertebrate lysozyme; MliC, membrane bound lysozyme inhibitor of c-type lysozyme), and one specific for g-type lysozyme (PliG, periplasmic lysozyme inhibitor of g-type lysozyme). Here, we investigated the role of these lysozyme inhibitors in virulence of Avian Pathogenic E. coli (APEC) using a serum resistance test and a subcutaneous chicken infection model. Knock-out of mliC caused a strong reduction in serum resistance and in in vivo virulence that could be fully restored by genetic complementation, whereas ivy and pliG could be knocked out without effect on serum resistance and virulence. This is the first in vivo evidence for the involvement of lysozyme inhibitors in bacterial virulence. Remarkably, the virulence of a ivy mliC double knock-out strain was restored to almost wild-type level, and this strain also had a substantial residual periplasmic lysozyme inhibitory activity that was higher than that of the single knock-out strains. This suggests the existence of an additional periplasmic lysozyme inhibitor in this strain, and indicates a regulatory interaction in the expression of the different inhibitors.     

An NB-LRR gene, <Emphasis Type="Italic">TYNBS1</Emphasis>, is responsible for resistance mediated by the <Emphasis Type="Italic">Ty</Emphasis>-<Emphasis Type="Italic">2 Begomovirus</Emphasis> resistance locus of tomato

Key message

An NB-LRR gene, TYNBS1, was isolated from Begomovirus-resistance locus Ty-2. Transgenic plant analysis revealed that TYNBS1 is a functional resistance gene. TYNBS1 is considered to be synonymous with Ty-2.

Abstract

Tomato yellow leaf curl disease caused by Tomato yellow leaf curl virus (TYLCV) is a serious threat to tomato (Solanum lycopersicum L.) production worldwide. A Begomovirus resistance gene, Ty-2, was introduced into cultivated tomato from Solanum habrochaites by interspecific crossing. To identify the Ty-2 gene, we performed genetic analysis. Identification of recombinant line 3701 confirmed the occurrence of a chromosome inversion in the Ty-2 region of the resistant haplotype. Genetic analysis revealed that the Ty-2 gene is linked to an introgression encompassing two markers, SL11_25_54277 and repeat A (approximately 200 kb). Genomic sequences of the upper and lower border of the inversion section of susceptible and resistant haplotypes were determined. Two nucleotide-binding domain and leucine-rich repeat-containing (NB-LRR) genes, TYNBS1 and TYNBS2, were identified around the upper and lower ends of the inversion section, respectively. TYNBS1 strictly co-segregated with TYLCV resistance, whereas TYNBS2 did not. Genetic introduction of genomic fragments containing the TYNBS1 gene into susceptible tomato plants conferred TYLCV resistance. These results demonstrate that TYNBS1 is a functional resistance gene for TYLCV, and is synonymous with the Ty-2 gene.

     

Detection of symbionts and virus in the whitefly <Emphasis Type="Italic">Bemisia tabaci</Emphasis> (Hemiptera: Aleyrodidae), vector of the <Emphasis Type="Italic">Mungbean yellow mosaic India virus</Emphasis> in Central India

Legume crops in Central India, the main soybean production area of the country, may suffer from yellow mosaic disease caused by the Mungbean yellow mosaic India virus (MYMIV). MYMIV is transmitted by the sweet potato whitefly, Bemisia tabaci (Gennadius), which is a species complex composed of various genetic groups. This vector species harbors different endosymbionts among regional strains and among individuals. To elucidate fundamental aspects of this virus vector in the state of Madhya Pradesh, the infection status of the symbionts and the virus in whiteflies was studied. A polymerase chain reaction (PCR) survey of the whiteflies collected in Madhya Pradesh found four secondary endosymbionts, Arsenophonus, Hemipteriphilus, Wolbachia, and Cardinium, in addition to the primary endosymbiont Portiera. Arsenophonus and Hemipteriphilus were highly infected but the infection rates of Wolbachia and Cardinium were low. MYMIV was detected in whitefly populations collected from various host plants in Madhya Pradesh. The whitefly populations belonged to the Asia I and II genetic groups; several different Asia II populations were also distributed. Specific relations were not observed among symbiont infection status, virus infection, and the whitefly genetic groups in the populations of Madhya Pradesh, though Cardinium was highly detected in the Asia II-1 group. New primers, which can be used for PCR template validation and for discriminating two phylogenetically close endosymbionts, were designed.