Epitope mapping of the Brucella melitensis BP26 immunogenic protein: usefulness for diagnosis of sheep brucellosis

Sequencing of bp26, the gene encoding the Brucella sp. immunogenic BP26 periplasmic protein, was performed in the reference strains of Brucella abortus, B. suis, and B. ovis. The three bp26 sequences were almost identical to that published for B. melitensis 16M bp26, and only minor nucleotide substitutions, without modifying the amino acid sequence, were observed between species. The bp26 genes of the seven B. abortus biovar reference strains and B. abortus S19 and RB51 vaccine strains were also sequenced. Again, only minor differences were found. Surprisingly, the bp26 nucleotide sequence for B. abortus S19 was almost identical to that found for B. melitensis 16M and differed from the sequence described previously by others (O. L. Rossetti, A. I. Arese, M. L. Boschiroli, and S. L. Cravero, J. Clin. Microbiol. 34:165-169, 1996) for the same B. abortus strain. The epitope mapping of BP26, performed by using a panel of monoclonal antibodies and recombinant DNA techniques, allowed the identification of an immunodominant region of the protein interesting for the diagnosis of B. melitensis and B. ovis infection in sheep. A recombinant fusion protein containing this region of BP26 reacted indeed, in Western blotting, as the entire recombinant BP26 against sera from B. melitensis- or B. ovis-infected sheep while it avoided false-positive reactions observed with sera from Brucella-free sheep when using the entire recombinant BP26. Thus, use of this recombinant fusion protein instead the entire recombinant BP26 could improve the specific serological diagnosis of B. melitensis or B. ovis infection in sheep.     

Analysis of myosin heavy chains at the protein level in horse skeletal muscle

Combined methodologies of enzyme-linked immunosorbent assay (ELISA), sodium dodecyl sulphate polyacrilamide gel electrophoresis (SDS-PAGE), immunoblotting, traditional myofibrillar ATPase (mATPase) histochemistry and immunocytochemistry of whole biopsied samples were used to study myosin heavy chain (MHC) isoforms in the equine gluteus medius muscle. The ELISA technique allowed the quantification of the three MHC isoforms known to be present in different horse muscles: slow (MHC-I) and two fast (termed MHC-IIA and MCH-IIX). The SDS-PAGE method resolved MHCs in three bands: MHC-I, MHC-IIX and MHC-IIA from the fastest to the slowest migrating band and a quantification by densitometry for each MHC isoform was also possible. The identity of these three MHCs was confirmed by immunoblots with specific monoclonal antibodies. Five fibre types were defined immunohistochemically according to their MHC content: I, I + IIA, IIA, the hybrid IIAX and IIX. When quantitative data obtained with the four different methodologies were combined and compared, they were consistent and, when considered together, showed significant correlation. Nevertheless, the percentage of MHC-IIA histochemically derived was underestimated, while that of MHC-IIX was overestimated in comparison with the immunocytochemical determination of these MHC isoforms. The percentage of MHC-I obtained by ELISA technique was underestimated. In short, these integrated methods for the analysis of MHCs at the protein level demonstrate that equine skeletal muscle does not express the MHC-IIB, so type II fibres have been misclassified in numerous previous studies based upon the very traditional mATPase histochemistry. They also offer new prospects for muscle fibre typing in equine experimental studies and veterinary medicine.     

Identification of Brucella melitensis vaccine strain Rev.1 by PCR-RFLP based on a mutation in the rpsL gene

The live attenuated strain B. melitensis Rev.1 is considered the best vaccine available for the prophylaxis of brucellosis in sheep and goats. The Rev.1 vaccine was obtained in the 1950s by a two-step selection involving firstly streptomycin resistance and dependence and secondly reversion of dependence but keeping streptomycin resistance. Chromosomally acquired streptomycin resistance is frequently due to mutations in the gene encoding the ribosomal protein S12, rpsL. Nucleotide sequencing revealed one mutation in the rpsL gene of vaccine strain Rev.1 compared to that of reference strain 16M leading to an amino acid Pro-to-Leu change at codon position 91 (Pro91Leu). This mutation resulted also in the lack of a NciI restriction site in the gene. PCR-restriction fragment length polymorphism (PCR-RFLP) using NciI applied to a large number of Brucella reference and field strains showed that the mutation detected was specific of vaccine strain Rev.1.     

Unusual parosteal osteochondromatous proliferation or Nora's tumor. A clinicopathological analysis of 4 cases

Nora's lesion, also known as bizarre parosteal osteochondromatous proliferation (B.P.O.P.), involves mostly the small tubular bones of the hands and feet. Histologically, it is characterized by a proliferation of chondroid, bony and fibrous tissues, sometimes with high cellular density, bizarre chondrocytes but is devoid of cellular atypia and necrosis. Distinct blue color is noted at the interface of bone and cartilage. The most important lesions that present differential diagnostic problems are chondrosarcoma, parosteal osteosarcoma and florid reactive periostitis. The lesion is benign but may recur locally in as many as 55%. The clinical and pathological findings of four cases of Nora's lesion are presented.     

Postmenopausal frontal fibrosing alopecia. Report of 3 cases

Postmenopausal frontal fibrosing alopecia is a rare aspect of scarring alopecia concerning elderly women. It appears as a receding anterior hair line localised in the frontal and temporal regions. It is a particular pathologic and clinical form of lichen planopilaris. The histologic aspect is that of a licheno?d inflammatory infiltrate affecting the dermal follicular junction, accompanied by a fibrous scarring aspect, the latter contributing to the diagnosis and individualization of this entity. Disco?d lupus erythematous is the main histologic differential diagnosis. Postmenopausal period is the only associated condition found in affected women. Evolution is unpredictable and does not seem to be modified by treatment.     

Lipopolysaccharide heterogeneity in Brucella strains isolated from marine mammals

Smooth lipopolysaccharides (S-LPSs) from Brucella strains isolated from seals, dolphins, porpoises, an otter and a minke whale were characterized by ELISA using monoclonal antibodies (mAbs) directed against seven previously defined O-polysaccharide (O-PS) epitopes and by Western blot after SDS-PAGE. All strains studied were A-dominant as shown by specific polyclonal sera and this was also confirmed by the mAbs. However, binding patterns in ELISA of mAbs to the specific common (C) epitopes were rather heterogeneous, and for some strains, such as those isolated from striped dolphins, binding of these mAbs was much reduced or negative as had previously been shown for Brucella suis biovar 2 strains. Western blot after SDS-PAGE showed the typical A-dominant strain banding pattern for all marine mammal Brucella isolates, but the average S-LPS size was shorter in many of these compared to reference Brucella abortus strain 544. Thus, S-LPSs of the marine mammal isolates show heterogeneity with regard to their O-PS C epitope content and their average size.     

Initial experience with two-incision laparoscopic adjustable gastric banding

BackgroundTo introduce the 2-incision technique for laparoscopic adjustable gastric banding (LAGB) and report our experience with 25 consecutive patients. Newer applications of minimally invasive laparoscopic techniques have been touted as revolutionary.MethodsWe have introduced a technique for LAGB that uses 2 skin incisions: 1 incision in the right upper quadrant (2.5 cm) that accommodates 2 trocars (11 and 5 mm) through which the dissection and implantation of the band were undertaken, and a 0.5-cm incision in the left upper quadrant for the 5-mm videoscope. The band reservoir was placed in a subcutaneous pocket through the upper quadrant incision. Previously, we used a standard 5-incision technique: 2 in the right upper quadrant, 2 in the left paramedian, and 1 in the subxyphoid area to retract the liver. The data from 25 consecutive 2-incision LAGB procedures (October 2007 to April 2008) were compared with the data from 19 consecutive standard 5-incision LAGB procedures (July 2007 to October 2007). The data are presented as mean ± SD. The t test was used to compare the mean values, and P <.05 was considered significant.ResultsThe mean estimated blood loss in the 2-incision LAGB was 54 ± 2 mL compared with 17 ± 1 mL in the standard technique (P = .040). The mean operating time for the 2-incision LAGB was 119 ± 1 minutes compared with 103 ± 1 minutes for the standard technique (P = .047). No mortality or procedure-related complications (e.g., erosion, slippage) occurred in the 2 groups.ConclusionTwo-incision LAGB is feasible; however, it is associated with an increased operating time and blood loss. The operating time and blood loss might improve with standardization of the operative technique and introduction of newly designed flexible tip instruments. Additional prospective studies with a larger sample size are needed to assess the efficacy and benefit of the 2-incision technique versus the standard technique.     

A cross‐sectional study of the catheter management of neurogenic bladder after traumatic spinal cord injury

Aims

This cross‐sectional study describes the catheter management of neurogenic bladder (NGB) in patients with traumatic spinal cord injury (tSCI) with emphasis on the motivations behind transitions between intermittent (IC) and indwelling catheters.

Methods

Patients at the Minneapolis VA with history of tSCI who utilized either intermittent catheterization (IC), urethral (UC) or suprapubic (SP) catheters, participated in a voluntary, anonymous survey regarding their bladder management strategies.

Results

A total of 100 patients participated, 94% were male and 90% Caucasian with median age of 61 years. Patients with current UC or SP were older than those utilizing IC (P = 0.002). The median age at injury and years since SCI were 32 years and 20.5 years, respectively. The median time with current modality was 11 years. A total of 27% of all patients reported at least one transition between catheter type. A total of 14 of 54 patients using IC had prior use of UC or SP, while 12/25 patients using SP and 10/21 patients using UC had prior use of IC. The most common reasons to stop IC included inconvenience, physician recommendation, and dislike of IC. A total of 53% of patients currently using UC or SP reported never using IC. Patients currently using SP were more content with their current catheterization method than those using UC or IC (P = 0.046).

Conclusions

Among patients using catheters for NGB, intermittent catheterization was the most common modality utilized and the transition between intermittent and indwelling catheter was most often influenced by patient preferences and clinician recommendations.