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Diaporthe species have often been reported as plant pathogens, endophytes and saprophytes, commonly isolated from a wide range of infected plant hosts. In the present study, twenty strains obtained from leaf spots of twelve host plants in Yunnan Province of China were isolated. Based on a combination of morphology, culture characteristics and multilocus sequence analysis of the rDNA internal transcribed spacer region (ITS), translation elongation factor 1-α (TEF), β-tubulin (TUB), calmodulin (CAL), and histone (HIS) genes, these strains were identified as eight new species: Diaporthe camelliae-sinensis, D. grandiflori, D. heliconiae, D. heterostemmatis, D. litchii, D. lutescens, D. melastomatis, D. pungensis and two previously described species, D. subclavata and D. tectonendophytica. This study showed high species diversity of Diaporthe in tropical rain forests and its hosts in south-western China.

Species of Diaporthe have often been reported as plant pathogens, endophytes or saprobes, commonly isolated from a wide range of plant hosts. Sixteen strains isolated from species of ten host genera in Yunnan Province, China, represented three new species of Diaporthe, D. chrysalidocarpi, D. machili and D. pometiae as well as five known species D. arecae, D. hongkongensis, D. middletonii, D. osmanthi and D. pandanicola. Morphological comparisons with known species and DNA-based phylogenies based on the analysis of a multigene (ITS, TUB, TEF, CAL and HIS) dataset support the establishment of the new species. This study reveals that a high species diversity of Diaporthe with wide host ranges occur in tropical rainforest in Yunnan Province, China.

Microdochium species have often been reported as plant pathogens and saprophytes and are commonly isolated from some diseased plant hosts. The primary aim of the present study was to describe and illustrate two new Microdochium species isolated from the leaf spot of Indocalamus longiauritus in Yunnan Province, China, namely Microdochium yunnanense and M. indocalami, spp. nov., based on their morphology and multilocus phylogenetic analyses of the combined ITS, LSU, TUB2, and RPB2. DNA sequence data indicate that six strains represent three independent groups from related and similar species in Microdochium. Microdochium indocalamisp. nov. clustered with M. fisheri, M. lycopodinum, M. rhopalostylidis, and M. phragmitis. Microdochium yunnanensesp. nov. grouped with M. bolleyi. In addition, the strain SAUCC1017 is recorded as an unidentified species in Microdochium. Descriptions and illustrations of the new species in the genus and Microdochium sp. indet. are provided.

Oxidative stress contributes a lot to initiation and progression of pathological conditions. Heme oxygenase 1 (HO1), a cytoprotective enzyme, is usually upregulated to alleviate oxidative stress in vivo. The function of teleost HO1 in the response to oxidative stress induced by heavy metal exposure and in pathogenic bacterial infection remains uncertain. In the present study, both complementary DNA and genomic sequence of a HO1-like gene cloned from the liver of swamp eel (Monopterus albus) are reported. Sequence analysis showed that the putative amino acid sequence contained a conserved heme oxygenase signature and displayed higher similarity to HO1 genes of other teleosts. Expression profile of swamp eel HO1 was investigated in healthy tissues and in tissues following stimulation with pathogenic bacteria (Aeromonas hydrophila) or cadmium chloride (CdCl2) exposure. Results demonstrated that HO1 messenger RNA (mRNA) was highly expressed in the liver and relatively less in other tissues. Bacterial infection with A. hydrophila significantly changed HO1 mRNA expression in the liver, spleen, and kidney, and the mRNA expression of HO1 and Nrf2 in the liver was elevated after the fish were exposed to CdCl2. Subsequently, the swamp eel HO1 was subcloned into a pET28a expression vector and transformed into Escherichia coli BL21 (DE3). Recombinant HO1 (rHO1) was successfully induced by 0.1 mmol/l IPTG and purified by Ni-NTA His Bind Resin purification system. To determine whether the rHO1 could confer stress tolerance in vitro, the viability of control and HO1-expressing E. coli under CdCl2 stress was compared by spot assay. The rHO1 protein significantly increased survival rates of the bacterial hosts. To evaluate whether intraperitoneal injection with rHO1 protected the liver of swamp eel against A. hydrophila-induced oxidative stress, mRNA expression of HO1, Nrf2, hepcidin, and IL-1β as well as the oxidative stress-related parameters (ROS and total antioxidant capacity (T-AOC)) in the liver were examined. The results showed that exogenous rHO1 could significantly upgrade the mRNA expression of HO1 and hepcidin, coupled with increased ROS and T-AOC levels. However, Nrf2 and IL-1β expression levels were significantly downregulated and upregulated, respectively. These results suggested that HO1 should not only play a protective role in oxidative stress response and its adverse effects deserved further investigation.

Asian swamp eel is a highly commercial fish, primarily for China and other Asian countries. The aim of this paper is to evaluate the genetic diversity of wild and cultured samples of Asian swamp eel Monopterus albus using ISSR markers. A total of 129 individuals belonging to three wild samples, Xiantao (XT), Huanggang (HG), Xinyang (XY) and three cultured samples, Wuhan (WH), Jingzhou (JZ) and Nanjing (NJ) were randomly selected for genetic analysis. Twelve ISSR primers were used for screening the six populations and 110 loci were obtained. The polymorphic loci were estimated to be 54%, 56.3%, 58.2%, 60.6%, 69.5% and 71% in NJ,WH, JZ, XT, HG and XY samples, respectively. Average heterozygosity value varied from 0.1956 to 0.2449. The three wild samples showed higher genetic diversity than the cultured samples (P < 0.05), including polymorphic bands (PPB), observed number of alleles per locus (to), effective number of alleles per locus (Ne), Nei’s gene diversity index (H) and Shannon’s information index (I).

Pectolytic enzymes are found mainly in fungi and bacteria. The most widely occurring enzymes are polygalacturonase (PGs), pectin methylesterase (PMEs) and pectate lyase (PLs) produced during the infection process and during culturing. The secretion of these enzymes results in the disorganization of the plant cell walls, which is responsible for the pathogenicity of the pathogens. These enzymes degrade the pectin of plants causing maceration of plant tissues and the enzyme activity increases under favourable environmental conditions. We have found that Phytophthora capsici, a pathogenic oomycete, produces levels of these three enzymes equal to those produced by soft-rotting Erwinia chrysanthemi. The activity of PGs, PLs and PMEs was investigated at the optimum temperature, pH and ionic strength in highly pathogenic P. capsici strains cultivated in two kinds of liquid medium containing either crude pepper extracts plus pectin or pectin as the carbon source. Virulence tests and enzymes activity showed that there was a high correlation between the enzyme activity and the pathogenicity of P. capsici. The effects of different carbon sources on the enzyme activity showed that pepper extract plus pectin was the best source for the carbon source.

Hepcidin is a cysteine-rich, dual-function peptide with antimicrobial activity that plays crucial roles in iron homeostasis. A few hepcidin-like genes have been isolated from teleost. Here, we have identified a hepcidin-like gene from rice field eel (RFE), Monopterus albus. Nucleotide sequences including cDNA and genomic DNA (GenBank accession numbers: FJ436808 and FJ594996, respectively) and deduced amino acid sequences were presented. In the 949 bp-long genomic sequence, two introns and three exons were identified. The full-length cDNA encodes a prepropeptide of 90 amino acid residues. RTPCR analysis suggested that hepcidin transcripts are highly abundant in the liver and kidney, less abundant in the heart, skin, brain, blood cells, intestine, spleen and stomach and undetectable in muscle. After challenged with Aeromonas hydrophila infection or iron-dextran stimulation, the hepcidin transcript levels were analyzed by RT-PCR. The results revealed that the expression of hepcidin dramatically increased at 24 h post-infection of the pathogen injection. Moreover, hepcidin mRNAs in the liver, intestine and brain were 2.4, 1.5 and 2-fold increase, respectively, compared with the control animals after 5 days in iron-dextran injected RFEs.Key words: Rice field eel, Monopterus albus, hepcidin, gene expression

Major histocompatibility complex (MHC) class II molecules play an important role in the immune response of vertebrates. In this paper, full-length MHC IIA cDNA was isolated from swamp eel (Monopterus albus) by rapid amplification of cDNA ends PCR. The genomic structure, molecular polymorphism, tissue distribution, and immune response of the MHC IIA gene to bacterial challenge were investigated. The full-length cDNA (GenBank accession No.: KC616308) is 1,509 bp in length including an 83 bp-long 5′ untranslated region (UTR) and a 709 bp-long 3′ UTR, which encoded a 238 amino acids protein. In the 2,339 bp-long MHC IIA genomic DNA, four exons and three introns were identified. Sequence comparison exhibited that the deduced amino acid sequence shared 27.1–66.3% identity with those of other species. Seven alleles were identified from five healthy individuals. Number of alleles per individual diversified from two to five. Five different 5′ UTR sequences and two different 3′ UTR sequences from one individual may infer the existence of five loci at least. Real-time quantitative PCR demonstrated that swamp eel MHC IIA transcripts were ubiquitously expressed in ten tissues, but the expression level was distinctly different. Significant changes were observed in liver, spleen, kidney and intestine after challenged with pathogenic bacteria Aeromonas hydrophilia.

Phytophthora capsici causes serious diseases in numerous crop plants. Polygalacturonases (PGs) are cell wall-degrading enzymes that play an important role in pathogenesis in straminopilous pathogens. To understand PGs as they relate to the virulence of P. capsici, Pcipg2 was identified from a genomic library of a highly virulent P. capsici strain. Pcipg2 was strongly expressed during symptom development after the inoculation of pepper leaves with P. capsici. The wild protein (PCIPGII) was obtained from the expression of pcipg2 and found that increasing activity of PGs in PCIPGII-treated pepper leaves was consistent with increasing symptom development. Asp residues in active sites within pcipg2 affected PCIPGII activity or its virulence on pepper leaves. Results show that pcipg2 is an important gene among pcipg genes, and illustrate the benefit of analyzing mechanisms of pathogenicity during the period of host/parasite interaction.