Lithium attenuated the depressant and anxiogenic effect of juvenile social stress through mitigating the negative impact of interlukin-1ß and nitric oxide on hypothalamic-pituitary-adrenal axis function.

The neuroimmune-endocrine dysfunction has been accepted as one of fundamental mechanisms contributing to the pathophysiology of psychiatric disorders including depression and anxiety. In this study, we aimed to evaluate the involvement of hypothalamic-pituitary-adrenal (HPA) axis, interleukin-1ß, and nitrergic system in mediating the negative behavioral impacts of juvenile social isolation stress (SIS) in male mice. We also investigated the possible protective effects of lithium on behavioral and neurochemical changes in socially isolated animals. Results showed that experiencing 4-weeks of juvenile SIS provoked depressive and anxiety-like behaviors that were associated with hyper responsiveness of HPA axis, upregulation of interleukin-1ß, and nitric oxide (NO) overproduction in the pre-frontal cortex and hippocampus. Administration of lithium (10 mg/kg) significantly attenuated the depressant and anxiogenic effects of SIS in behavioral tests. Lithium also restored the negative effects of SIS on cortical and hippocampal interleukin-1ß and NO as well as HPA axis deregulation. Unlike the neutralizing effects of l-arginine (NO precursor), administration of l-NAME (3 mg/kg) and aminoguanidine (20 mg/kg) potentiated the positive effects of lithium on the behavioral and neurochemical profile of isolated mice. In conclusion, our results revealed that juvenile SIS-induced behavioral deficits are associated with abnormalities in HPA-immune function. Also, we suggest that alleviating effects of lithium on behavioral profile of isolated mice may be partly mediated by mitigating the negative impact of NO on HPA-immune function.

Expression analysis of microRNA-125 in patients with polycythemia vera and essential thrombocythemia and correlation with JAK2 allele burden and laboratory findings.

INTRODUCTION: The JAK2V617F mutation has emerged in recent years as a diagnostic as well as a treatment target in patients with polycythemia vera (PV) and essential thrombocythemia (ET). The disease phenotype is also influenced by other factors such as microRNA (miRNA) deregulation. The aim of this study was to investigate miR-125 expression level in these patients with those obtained from healthy control subjects and its correlation with JAK2 allele burden and laboratory findings. METHODS: In total, forty patients with a clinical diagnosis of PV and ET were examined at the time of diagnosis. Ten healthy subjects were checked as controls. We performed JAK2 V617F allele burdens measurement and expression analysis of miR-125b-5p, miR-125b-3p, miR-125a-5p, and miR-125a-3p in leukocytes isolated from peripheral blood by quantitative real-time polymerase chain reaction. RESULTS: MiR-125b-5p and miR-125a-5p were upregulated in both patients with PV (P = 0.00 and P = 0.003, respectively) and ET (P = 0.02 and P = 0.002, respectively). In PV group, a significant correlation was observed between miR-125a-5p and platelet counts (P = 0.01, r = 0.531). The correlation between miRNA and JAk2 allele burden was not significant. CONCLUSION: In conclusion, our data indicate that other factors such as aberrant miR-125 expression may influence on the disease phenotype in patients with PV and ET.

Apigenin modulates the expression levels of pro-inflammatory mediators to reduce the human insulin amyloid-induced oxidant damages in SK-N-MC cells.

Amyloid depositions of proteins play crucial roles in a wide variety of degenerative disorders called amyloidosis. Although the exact mechanisms involved in amyloid-mediated cytotoxicity remain unknown, increased formation of reactive oxygen species (ROS) and nitrogen species and overproduction of pro-inflammatory cytokines are believed to play key roles in the process. In that regard, we investigated the effect of apigenin, a common dietary flavonoid with high antioxidant and anti-inflammatory properties on potential factors involved in cytotoxicity of human insulin amyloids. Pretreatment of SK-N-MC neuroblastoma cells with apigenin increased cell viability and reduced the apoptosis induced by insulin fibrils. In addition, apigenin attenuated insulin fibril-induced ROS production and lipid peroxidation. Our result also demonstrated that pretreatment of the fibril-affected cells with apigenin caused an increase in catalase activity and the intracellular glutathione content along with reduction in nitric oxide production and nuclear factor κB, tumor necrosis factor α, and interleukin 6 gene expression based on real-time polymerase chain reaction evaluation. In accordance with these results, apigenin could be a promising candidate in the design of natural-based drugs for treatment or prevention of amyloid-related disorders.

The influence of the HLA-DRB1 and HLA-DQB1 allele heterogeneity on disease risk and severity in Iranian patients with multiple sclerosis.

Multiple sclerosis (MS) is a common autoimmune disorder of the central nervous system. Recent studies have shown that the HLA-DRB1 and DQB1 alleles are associated with MS susceptibility and severity. However, this is controversial in different population studies. In the present study, the roles of HLA-DRB1 and DQB1 alleles and the amino acids were investigated on disease risk and severity in 120 Iranian patients with MS and 120 controls. Our findings indicate that the DRB1*1501 allele (OR = 3.203 P = 0.001), the DRB1*1501-DQB1*0602 haplotype (OR = 7.792 P = 0.003) and the DRB1*1501/0701- genotype (OR = 3.320 P = 0.006) and amino acid Leu26 (OR = 1.645 P = 0.005) and Phe9 (OR = 1.893 P = 0.009) on the DQß1 chain are significantly associated with MS susceptibility. DRB1*1001 was the only allele that had a protective effect against MS (P = 0.0004). We also found that the DQB1*0303 allele was significantly associated with disease severity (mean Multiple Sclerosis Severity Score difference = 1.979, P = 0.002). However, protective effect of the DRB1*1001 against MS and also association of DQB1*0303 allele with MS severity need to be confirmed by larger sample size.

Direct short-term cytotoxic effects of BIBR 1532 on acute promyelocytic leukemia cells through induction of p21 coupled with downregulation of c-Myc and hTERT transcription.

Acute promyelocytic leukemia (APL) is characterized by specific t(15;17), distinct morphologic picture, and clinical coagulopathy that contribute to the morbidity and mortality of the disease. This study aims to investigate the effects of antitelomerase compound BIBR1532 on APL cells (NB4). BIBR 1532 exerts a direct short-term growth suppressive effect in a concentration-dependent manner probably through downregulation of c-Myc and hTERT expression. Our results also suggest that induction of p21 and subsequent disturbance of Bax/Bcl-2 balanced ratio as well as decreased telomerase activity may be rational mechanisms for the potent/direct short-term cytotoxicity of high doses of BIBR1532 against NB4 cells.

The detection of disseminated tumor cells in bone marrow and peripheral blood of gastric cancer patients by multimarker (CEA, CK20, TFF1 and MUC2) quantitative real-time PCR.

OBJECTIVE: To investigate the suitability of multimarker detection of DTCs in PB and BM of GC patients. DESIGN AND METHOD: A qRT-PCR assay was developed to estimate the number of CEA, CK20, TFF1 and MUC2 transcripts in PB and BM samples of 35 GC patients prior to the initiation of therapy. PB samples from healthy volunteers and BM from patients with hematological malignancies were used as negative controls. RESULTS: In PB analysis; 22.9%, 37.1%, 31.4%, and 22.9% of GC patients and in BM analysis; 20%, 28.6%, 45.7%, and 22.9% of GC patients were positive for CEA, CK20, TFF1 and MUC2 mRNAs, respectively. Samples from the control group were negative for the expression of all the markers tested in this study. A higher positive ratio was obtained with the multimarker detection in comparison to the single marker detection. There was a significant correlation between the PB and BM samples for DTC detection. CONCLUSION: Multimarker detection assay is a reliable and powerful tool for the early detection of DTCs in GC patients.

The expression of p38, ERK1 and Bax proteins has increased during the treatment of newly diagnosed acute promyelocytic leukemia with arsenic trioxide.

BACKGROUND: Promising reports exist regarding the use of arsenic trioxide (ATO) as first-line treatment in acute promyelocytic leukemia (APL). Although the in vitro effect of ATO is extensively studied, the in vivo mechanism(s) of ATO action is mostly unknown. PATIENTS AND METHODS: Newly diagnosed APL patients were involved and received ATO (0.15 mg.kg/day) for 28 days as induction followed by consolidation therapy. Bone marrow (BM) aspirates were obtained on days 0, 14 and 28 of treatment for further molecular studies. Clinical findings and white blood cell counts were recorded as well. RESULTS: Complete remission was observed in 17 (85%) patients with the median duration of 28 days (18-38) and cumulative dosage of median 280 mg (180-350). Hyperleukocytosis and APL differentiation syndrome (63%), gastrointestinal disorders (30%), liver enzyme elevation and night sweating (50%) were the most prevalent side-effects. The expression of Bax, ERK1 and p38 proteins and caspase-3 activity increased significantly in promyelocytes of BM aspirates at days 14 and 28 of induction therapy. CONCLUSION(S): These findings point toward the role of p38 and Bax in the induction of apoptosis, which was confirmed by increase in caspase-3 activity. However, the increase in ERK1 expression with regard to leukocytosis could translate to a proliferative/differentiation effect.

Hepatocytes of donor origin in recipient liver after hematopoietic SCT in beta-thalassemia major patients.

BM and circulating cells contain stem cells with the potential to differentiate into mature cells of various organs. We determined whether stem cells transformed into hepatocytes. Biopsy specimens from liver were obtained from 11 patients who had undergone transplantation of hematopoietic stem cells from peripheral blood (eight patients) or BM (three patients). Four female patients had received transplants from a male donor and seven male patients had received transplants from a female donor. All patients had beta-thalassemia major and fibrosis in biopsy specimens from the liver before hematopoietic SCT. Hematopoietic stem cell engraftment was verified by STR analysis. The biopsies were studied for the presence of donor-derived hepatocytes using FISH of interphase nuclei and immunohistochemical staining for CD45 (leukocyte common Ag), and a hepatocyte-specific Ag. All 11 recipients of sex-mismatched transplants showed evidence of complete hematopoietic donor chimerism. XY-positive hepatocytes accounted for 4-6.7% of cells in histological sections of the biopsy specimens of female patients and XX-positive hepatocytes accounted for 3-7% of cells in histological sections of the biopsy specimens of male patients. These cells were detected in liver tissue as early as 1 year and as late as 8.5 years after hematopoietic SCT. BM and circulating stem cells can differentiate into mature hepatocytes in beta-thalassemia major patients who had undergone hematopoietic SCT.

Survey of factors effective on re-intubation among children admitted to pediatric intensive care unit.

This study was aimed to recognize the risk factors of re intubation among children who were admitted to pediatric intensive care unit. in an analytical cross-sectional study, the risk factors of reintubation in two groups of patients compared, both groups consist of 55 children, one with successful extubation and another with extubation failure. The study showed that neuromuscular disorders are the main underlying disease in extubation-failure group (p = 0.004). Besides, in comparison between two group of patients who had successful versus failed extubation, hypercapnia (PaCO2 > 50 mmHg) was shown to be the most common cause of both the first intubation (p = 0.003) and reintubation (p = 0.002) in patients who failed extubation. This study shows that neuromuscular disorders as a background, are the most common causes which defeat weaning from ventilator or result in reintubation by induction of hypercapnia.

Telomerase activity and telomere length in patients with acute promyelocytic leukemia: indicative of proliferative activity, disease progression, and overall survival.

BACKGROUND: The progressive shortening of telomeres and the activation of telomerase have been considered to be one of the key mechanisms in cellular immortalization and tumor progression. PATIENTS AND METHODS: About 300 sequential samples were collected from 40 patients during the course of acute promyelocytic leukemia (APL) disease. Telomerase activity (TA) and terminal restriction fragment (TRF) length were assessed by TRAP and Southern blot analyses, respectively. PML-retinoic acid receptor alpha (RARa)/glucose-6-phosphate dehydrogenase transcripts were quantified by real-time PCR. RESULTS: About 90% of the patients had a significant reduction in telomere length (TL) relative to the control (median 3.5 versus 11.37 kbp; P < 0.001). A significant positive correlation between TL and PML-RARa expression was found (P = 0.001). Telomerase was activated in all patients; however, TA level was significantly higher in the group of relapsed patients than patient with newly diagnosed. The group of patients with shortened TRF and elevated TA had a significantly poorer overall survival. CONCLUSIONS: The shortened TL and elevated TA in APL patients are mainly indicative of extensive proliferative activity and they correlate with disease progression and relapse; thus, they may serve as prognostic factors for a subset of APL patients with more aggressive disease and poor outcome, those who may not respond favorably to arsenic therapy.

Fludarabine and busulfan as a myeloablative conditioning regimen for allogeneic stem cell transplantation in high- and standard-risk leukemic patients.

Busulfan and cyclophosphamide (BuCy) are currently the most widely used myeloablative regimen to treat malignancies with allogeneic stem cell transplantation. Fludarabine has considerable efficacy in both immunosuppression and tumor cells killing with a minimal extramedullary toxicity. We evaluated the efficacy of 40 mg/m(2) fludarabine i.v. for 5 days and busulfan 4 mg/kg/day p.o. for 4 days as myeloablative conditioning regimen in 70 patients (median age 24 years) with acute leukemia or chronic phase of myelogenous leukemia. They all had human leukocyte antigen-matched sibling donors. The patients received 10 mug/kg granulocyte colony stimulating factor (GCSF), 24 h after stem cell infusion until engraftment occurred. Graft-versus-host disease (GVHD) prophylaxis included 3 mg/kg cyclosporine-A i.v. from day -2 to +6 followed by 12 mg/kg p.o. until day +60. The median time of neutrophil recovery (>0.5 x 109/l) and platelet recovery (>20 x 109/l) were 10 and 12 days, respectively. Mucositis (93%) and hepatic toxicity (16%) resolved with conservative therapy. The incidence of acute GVHD grade I-II and III-IV were 38.6 and 15.7% respectively. Overall survival and disease-free survival were 71 and 64% respectively with 17 months median follow-up for surviving patients. We conclude that FluBu may be used as a substitute for BuCy with almost the same efficacy and with a lower transplant adverse effect but to increase anti-leukemic effects, especially in acute lymphoblastic leukemia patients, it needs some modifications.

Real-time PCR analysis of PML-RAR alpha in newly diagnosed acute promyelocytic leukaemia patients treated with arsenic trioxide as a front-line therapy.

BACKGROUND: Recently, patients with acute promyelocytic leukaemia (APL) have experienced significant clinical gains after treatment with arsenic trioxide. However, the use of this agent as a front-line therapy for newly diagnosed patients is unclear. PATIENTS AND METHODS: Of 95 newly diagnosed APL patients, 85 patients who achieved complete remission (CR) were sequentially evaluated during a 4-60 month period by conventional RT-PCR. A total of 30 patients (six relapsed and 24 in continued CR) were selected and monitored by quantitative real-time PCR (RQ-PCR) assay. The PML-RARalpha fusion transcripts values were normalised to every 10(6) copies of G6PDH transcripts (NQ). RESULTS: RQ-PCR analyses showed a rapid rate of clearance of NQ levels during the courses of arsenic therapy. In the majority of patients in CR, the NQ levels were below 5 x 10(2) in peripheral blood (PB) samples. In all the relapsed cases with follow-up intervals of 1-6 months (median 3 months) clinical relapse was predictable by increasing NQ level above this threshold. CONCLUSIONS: Our study highlights the usefulness of PB and the definition of threshold level for early prediction of relapse. The threshold level correlates well with risk of relapse; therefore, transcript ratio below the level should be regarded as a goal in the clinical management of this disease.

Treatment of acute promyelocytic leukemia with arsenic trioxide without ATRA and/or chemotherapy.

INTRODUCTION: Arsenic trioxide is effective and approved for treatment of relapsed or refractory acute promyelocytic leukemia (APL) cases resistant to all-trans retinoic acid (ATRA), but its effect on new cases of APL is not clear. MATERIALS AND METHODS: We studied 111 patients with APL. Arsenic trioxide was infused at 0.15 mg/kg daily dose, until complete remission was achieved. Then, after 28 days of rest, arsenic trioxide was infused daily for 28 days as consolidation therapy. We studied minimal residual disease (MRD) by semi-sensitive reverse transcription polymerase chain reaction (RT-PCR) on peripheral blood samples. RESULTS: Complete remission was observed in 95 patients (85.6%). With the median (range) follow-up period of 16.5 (1-57) months, 1- and 2-year disease-free survival was 88.3% and 63.7%, respectively; 24 patients relapsed, 19 of whom achieved a second complete remission, again by arsenic trioxide. Third and fourth remissions were seen in some relapsed patients, again by arsenic trioxide. For patients in complete remission, 1- and 3-year survival was 95.5% and 87.6%, respectively. MRD was positive in four (8.3%) out of 48 cases during 1 year after remission induction; three of them relapsed clinically. CONCLUSIONS: Arsenic trioxide is effective as first-line treatment for APL. Results of arsenic trioxide combination therapy with chemotherapy/ATRA requires further study.

Structure and genomic organization of a second cluster of immunoglobulin heavy chain gene segments in the channel catfish.

The structure, organization, and partial sequence of a 25-kb genomic region containing a second cluster of H chain gene segments in the channel catfish (Ictalurus punctatus) has been determined. Multiple VH gene segments, representing different VH families, are located upstream of a germline-joined VDJ. The VDJ segment has a split leader sequence and a single open reading consistent with that expressed in members of the VH1 family. Downstream of the germline-joined VDJ is a single JH segment and two pseudogene exons structurally similar to the Cmu1 and Cmu2 exons of the functional gene. Both pseudogene exons are multiply crippled with RNA splice sites destroyed, and open reading frames are interrupted by termination codons, insertions, and/or deletions. Sequence alignment of a 10.8-kb region within the second H chain cluster with the genomic sequence of the nine JH segments and the functional Cmu within the first H chain gene cluster indicates that the second H chain gene cluster probably arose by a massive duplication event. The JH region of the VDJ, the coding and flanking regions of the single JH segment, and the pseudogene Cmu exons were readily aligned with homologous segments in the first gene cluster. This duplication event may have extended to include the upstream VH segments. A member of the Tc1 mariner family of transposable elements is located downstream of the pseudogene Cmu2, which suggests that the transposition may have contributed to the evolution of the duplicated Cmu.

Structure and genomic organization of a second class of immunoglobulin light chain genes in the channel catfish.

Earlier studies distinguished two classes of catfish light (L) chain (designated F and G). The cDNA structure and genomic organization of G L chain gene clusters has also been characterized previously. In this study, full length cDNA encoding F L chain was derived using PCR strategies based on the determined amino-terminal protein sequence. The encoded V region is readily delineated into framework regions (FR) and complementarity-determining regions (CDR). Multiple sequence alignments indicate that the F V(L) is closely related to kappa gene families. The F C(L) cannot be generally classified but it is structurally distinct from the C(L) regions of G: the amino acid sequence similarity is <35%. cDNA sequences representing processed sterile F transcripts of different loci were identified. Each sequence begins within the J(L) recombination signal sequence and extends downstream through the I(L)-C(L) segments. Genomic blots hybridized with C(L) probes indicate that there are at least 50 different C(L) segments. Based upon V(L) hybridization studies, different families of V(L) segments appear to be associated with closely related F C(L) segments. In characterized genomic clones, F gene segments are arranged in closely linked clusters with single copies of V(L), J(L), and C(L) segments within each cluster. The V(L) segments are located in opposite transcriptional polarity relative to the J(L) and C(L) segments, which indicates that V(L) segments rearrange by inversion. These combined studies establish that two structurally distinct classes of L chains are present in teleost fish and that both of the L chain classes evolved within a common organizational pattern of clustered segmental genes.

Structure and genomic organization of VH gene segments in the channel catfish: members of different VH gene families are interspersed and closely linked.

To determine the structure and organization of germline VH gene segments in the channel catfish, genomic lambda libraries were screened with cDNA probes representing different catfish VH gene families. Thirty-six VH positive genomic clones were isolated and four of these were characterized by restriction mapping and Southern blot analysis with probes specific for each known VH gene family. The four clones, representing about 65 kb of DNA, contained 21 VH segments. The average distance between segments was about 3 kb and gene segments representing different VH gene families were interspersed with each other. Dot-blot hybridization analysis of all 36 genomic clones (average insert size 16-18 kb) indicated that the average clone contained gene segments representing four different VH families. In addition, these analyses indicated that VH segments representing each VH family could be found closely linked to gene segments representing each of the other VH families. Genomic restriction fragments containing a VH segment of each gene family were sequenced. These analyses showed that the general structure of VH segments is conserved in catfish. These structural features include the presence of a leader sequence split by a short intron, an uninterrupted open reading frame encoding readily identified framework and complementarity determining regions, and a downstream recombination signal sequence represented by a consensus heptamer, a 22-24 bp spacer, and an A-rich nonamer. Upstream of the VH segments was an octamer sequence. These analyses indicate that the organization and structure of VH segments typically associated with VH loci of higher vertebrates evolved early in phylogeny at the level of the bony fishes.

Structure and genomic organization of immunoglobulin light chain in the channel catfish. An unusual genomic organizational pattern of segmental genes.

Channel catfish L chain cDNA was obtained through a PCR strategy and used to isolate multiple L chain clones from cDNA and genomic libraries. Sequence analysis of full-length cDNA indicates that the V region is preceded by a leader peptide, and represented by framework and CDR regions. Both VL and CL domains contain the invariant cysteines and tryptophans as well as other phylogenetically conserved L chain residues. The sequence similarity of the catfish L chain with higher vertebrate kappa- and lambda-chains, however, does not readily allow the catfish L chain to be classified. Eight cDNA clones isolated from a cDNA library were shown to represent different processed derivatives of sterile L chain transcripts. These transcripts share a similar upstream sequence region and extend downstream to include a CL or alternatively a JL segment in partial germ-line configuration that has been spliced into a CL. Sequence comparisons indicate that these transcripts represent the product of different L chain loci. Genomic Southern blot analyses with VL and CL probes indicate that there are at least 30 VL segments and at least 15 CL segments. The analysis of 17 genomic L chain clones showed that each hybridized with VL-, JL-, and CL-specific probes. Characterization of the gene segments in three of these clones indicates a previously undescribed pattern of segmental gene organization. Gene segments are found in clusters with VL, JL, and CL segments in each cluster. Within a cluster VL segments reside upstream of single copies of closely linked JL and CL segments. The proximity of VL segments downstream from JL-CL segments suggests that individual clusters may be closely linked. The VL segments are located in opposite transcriptional polarity relative to the JL and CL gene segments, which indicates that VL segments are likely rearranged to JL-CL segments by inversion rather than deletion events.

Heavy chain joining region segments of the channel catfish. Genomic organization and phylogenetic implications.

The JH locus of the channel catfish has been characterized to determine the organization and structural diversity of JH segments. These analyses indicate that there are a total of nine JH segments tightly clustered within a region spanning about 2.2 kb. The JH locus is closely linked to the CH 1 domain of the expressed catfish H chain; the distance between the CH proximal JH segment (JH9) and the CH 1 domain is about 1.8 kb. Each JH segment has an upstream recombination sequence, which includes a T-rich nonamer, a 22- to 24-bp spacer, and a phylogenetically conserved heptamer. Each JH segment also has an open reading frame that encodes the conserved framework region 4 tryptophan (Trp-103) and terminates with a RNA donor splice site. The catfish JH locus contains an internal repetitive sequence region characterized by a short (183-188 bp) repeat that occurs sequentially five times. Strong sequence homology as well as the unified length of the repeated sequences indicate that JH segments JH3-JH7 probably arose as the result of a series of homologous but unequal crossover events. Sequence alignments of the duplicated JH segments indicates that there is diversity within the 5-11 nucleotides located immediately downstream from the heptamer, an observation which indicates that closely related JH segments can serve to enhance CDR3 diversity in the expressed H chain. Comparisons of the genomic JH sequences with different cDNA clones indicate that each JH segment is probably functional and that junctional diversity serves an important role in the generation of CDR3 diversity. In addition, single base differences observed in comparisons of JH-encoded regions indicate that there is probably somatic mutation or allelic variation of genomic JH segments. These studies suggest that the characteristic structure and organizational pattern of JH segments in higher vertebrates may have evolved early in vertebrate phylogeny at the level of the bony fish.

Plasmid and serological differences between Edwardsiella ictaluri strains.

Several studies have shown that isolates of Edwardsiella ictaluri obtained from infected channel catfish in the southeastern United States harbor two cryptic plasmids, designated pCL1 (5.7 kb) and pCL2 (4.9 kb). These isolates appear to be serologically homogeneous. To extend these studies, we focused our analyses on two isolates of nonictalurid origin. Plasmid analyses of a danio isolate showed that it harbored plasmids which were similar if not identical to pCL1 and pCL2. This strain was also serologically indistinguishable from those isolated from channel catfish. In contrast, a green knife fish (GNF) isolate harbored four plasmids with relative mobilities of 6.0, 5.7, 4.1, and 3.1 kb. Southern blot analyses indicated that only the 5.7- and 4.1-kb plasmids strongly hybridized under high-stringency conditions to probes specific for pCL1 and pCL2, respectively. The GNF isolate showed minimal reactivity when reacted with polyclonal antiserum prepared against a channel catfish isolate. However, polyclonal antiserum to the GNF isolate strongly reacted with the GNF isolate in both surface fluorescence and agglutination reactions. Sodium dodecyl sulfate-polyacrylamide gel electrophoresis analyses of cell lysates showed that the protein banding patterns of the strains compared were similar. However, Western blots of proteinase K-digested cell extracts showed that O antigen of the GNF isolate was antigenically distinct from the O antigen of the other isolates. These studies indicate that there are different serotypes of E. ictaluri and suggest that plasmid and serological analyses of future isolates of E. ictaluri can be used to determine whether structurally distinct strains are emerging in major channel catfish aquaculture areas.

Immunoglobulin heavy chain constant and heavy chain variable region genes in phylogenetically diverse species of bony fish.

Genomic DNA from 18 phylogenetically diverse species of bony fish was hybridized with probes specific for the channel catfish immunoglobulin heavy chain constant (CH) gene, as well as with immunoglobulin heavy chain variable (VH) probes specific for five channel catfish VH gene families. The results showed that CH probes strongly hybridized only to genomic fragments from other catfish species. In contrast, restricted DNA from most other species hybridized with at least two channel catfish VH probes. In those species whose DNA hybridized with multiple VH probes, the restriction pattern of hybridizing fragments was probe-dependent. These studies suggest that (1) the CH gene defined in channel catfish appears to share similarity only with CH genes in other catfish species, (2) families of VH genes appear to have diverged in early phylogenetic lineages of teleosts, and (3) VH genes similar to those defined in catfish appear to be widely represented in phylogenetically diverse species of teleosts.