Insight into the Machinery and Applications for Understanding the Pathogen- Vector Interface

The availability of genome sequencing data in combination with knowledge of expressed genes via transcriptome and proteome data has greatly advanced our understanding of arthropod vectors of disease. Not only have we gained insight into vector biology, but also into their respective vector-pathogen interactions. By combining the strengths of postgenomic databases and reverse genetic approaches such as RNAi, the numbers of available drug and vaccine targets, as well as number of transgenes for subsequent transgenic or paratransgenic approaches, have expanded. These are now paving the way for in- field control strategies of vectors and their pathogens. Basic scientific questions, such as understanding the basic components of the vector RNAi machinery, is vital, as this allows for the transfer of basic RNAi machinery components into RNAi-deficient vectors, thereby expanding the genetic toolbox of these RNAi-deficient vectors and pathogens. In this review, we focus on the current knowledge of arthropod vector RNAi machinery and the impact of RNAi on understanding vector biology and vector-pathogen interactions for which vector genomic data is available on VectorBase

Functional outcomes and quality of life following complex tibial fractures treated with circular external fixation : a comparison between proximal, midshaft, and distal tibial fractures

AIM AND OBJECTIVE: The purpose of this study was to compare clinical results following complex proximal, midshaft, and distal tibial fractures and investigate whether there are differences in outcomes between these locations. MATERIALS AND METHODS: Patients between 18 years and 65 years of age and minimum follow-up of 12 months with complex tibial fractures treated with a circular ring fixator were included. Functional outcomes were assessed using the Association for the Study and Application of Methods of Ilizarov (ASAMI) functional and bone scores, Foot Function Index (FFI), Four Step Square Test (FSST), and Timed Up and Go Test (TUG). Quality of life was assessed by the EQ-5D score. RESULTS: A total of 45 patients were included: proximal fractures, n= 11; midshaft fractures, n= 17; and distal fractures, n= 17. ASAMI functional (p= 0.8) and bone scores (p= 0.3) were not different. Excellent and good bone scores were achieved in >90% in all groups. FFI was 30.9 + 24.7 in the proximal group, 33.9 + 27.7 in the midshaft group, and 28.8 + 26.9 in the distal group (p = 0.8). TUG was 9.0 + 2.7 sec in the proximal group, 9.0+3.5 in the midshaft group, and 8.5+2.0 in the distal group (p = 0.67). FSST was 10.7 + 2.5 sec in the proximal, 10.3 + 3.8 in the midshaft, and 8.9 + 1.8 in the distal fracture groups (p = 0.5). EQ-5D index value was highest in the distal (0.72), lowest in the proximal (0.55), and 0.70 in the midshaft fracture groups (p = 0.001). EQ-5D VAS was significantly different between the proximal (65) and midshaft (82.3) (p = 0.001) and between the distal (75) and proximal (65) fracture groups (p = 0.001). CONCLUSIONS: The results of this study suggest that the functional outcomes between proximal, midshaft, and distal complex tibial fractures are comparable. Their ability to ambulate afterward is comparable to age-related normative data, but complex tasks are more difficult and better compared to the ambulating ability of a healthy population aged 65 to 80 years. Patients with proximal tibial fractures had significantly more disability by at least one functional level and/or one health dimension.https://www.stlrjournal.com/journalDetails/STLRdm2022Orthopaedic Surger

Expression profiling, gene silencing and transcriptional networking of metzincin metalloproteases in the cattle tick, Rhipicephalus (Boophilus) microplus

Tick proteins functioning in vital physiological processes such as blood meal uptake, digestion and reproduction are potential targets for anti-tick vaccines, since vaccination could inhibit these essential functions and ultimately affect tick survival. In this study we identified metzincin metalloproteases from Rhipicephalus microplus as potential vaccine candidates since they are implicated as essential to blood-cavity formation, bloodmeal digestion and reproduction in ixodid ticks. Eight transcripts encoding proteins that contain the characteristic metzincin zinc-binding motif HEXXHXXG/NXXH/D and a unique methionine containing “methionine-turn” were identified from native and in-house assembled R. microplus expressed eequence tag (EST) databases. These were representative of five reprolysin-like and three astacin-like metzincin metalloproteases. Reverse transcription-PCR analysis indicated that the reprolysins were most abundantly expressed in the salivary glands, whereas the astacins were most abundant in the midgut and ovaries. In vivo gene silencing was performed to assess a possible phenotype of these metalloproteases during adult female R. microplus blood feeding and reproduction. RNA interference (RNAi) against two of the reprolysins and one of the astacins significantly affected the average egg weight and oviposition rate. Evidently, this reverse genetic approach enabled the evaluation of the overall vital impact of tick proteins. Finally, integrated real time-PCR studies also revealed an extensive cross organ network between the R. microplus metzincin transcripts, supporting the use of a combinatorial metzincinbased anti- R. microplus vaccine.Research on anti-tick vaccines is supported by the Wellcome Trust under the ‘Animal Health in the Developing World’ initiative through project 075799 entitled ‘Adapting recombinant anti-tick vaccines to livestock in Africa’. International research was funded by the University of Pretoria’s ‘Postgraduate study abroad bursary program’.http://www.elsevier.com/locate/vetparab201

A reverse genetics approach to evaluate Metzincins as anti-Rhipicephalus microplus tick vaccine candidates

Tick proteins functioning in vital physiological processes such as blood meal uptake, digestion and reproduction are potential targets for anti-tick vaccines, since vaccination could disrupt these essential functions and ultimately affect tick survival. In this study we identified metzincin metalloproteases from R. microplus, the world’s most economically important external ectoparasite of cattle, as potential vaccine candidates since they are implicated to be essential to blood-cavity formation, bloodmeal digestion and reproduction in ixodid ticks. A vaccine derived from a single member of such a large family pose the obstacle of redundancy within the family, that may allow the function of the targeted family member to be taken up by other family members. Therefore the aim of this study was not only to focus on the physiological importance of each metzincin transcript, but also to investigate the differential gene expression network between the different metzincin family members. Eight transcripts encoding proteins containing the characteristic metzincin zinc-binding motif HEXXHXXG/NXXH/D and a unique methionine-turn were identified from native and in-house assembled R. microplus Expressed Sequence Tag databases. These were representative of five reprolysin-like and three astacin-like metzincin metalloproteases. Reverse transcription-PCR indicated that the reprolysins were most abundantly expressed in the salivary glands, whereas the astacins were most abundant in the midgut and ovaries. In vivo gene silencing utilizing RNA interference, was performed to assess a possible phenotype in silenced adult female R. microplus ticks during blood feeding and reproduction. RNAi against two reprolysins and one astacin significantly affected average egg weight as well as the oviposition rate. Moreover, integrated real time-PCR studies revealed an extensive cross organ network between the R. microplus metzincin transcripts, supporting the use of a combinatorial metzincinbased anti- R. microplus vaccine targeting multiple members of the large metzincin clan simultaneous. To conclusively evaluate the vaccination potential of the three identified metzincin candidates, the immunogenicity and protective properties of the recombinant proteins needs to be determined. Due to metalloproteases destructive activity and characteristics such as cysteine rich domains, only selected domains of the three candidates were expressed, using a cost effective Eshericia coli based expression system. Finally, the ability of each successfully expressed domain to elicit an immune response and serve as a protective antigen against R. microplus will be screened during vaccination trials in cattle.Thesis (PhD)--University of Pretoria, 2013.gm2013Biochemistryunrestricte

The tibial bayonet method of wound closure

Abstract Management of open lower limb fractures with soft tissue defects can be a technically challenging orthopaedic problem. Limited availability of orthoplastic services means that alternatives to the fix and flap concept are required in order to prevent infected non-unions from developing. The proposed ‘bayonet apposition’ allows the surgeon to temporarily shorten the limb without angulating the limb or creating a bone defect and removing viable bone. The viable bone edges are overlapped in a bayonet-like manner in order to appose the wound and skin edges. The limb length is restored by gradually distracting the bone segments once the soft tissues have healed. This is facilitated with a hexapod fixator for stabilization of the fracture and distraction. Prerequisites for utilizing this method are circumferential soft tissue damage to the lower limb with viable distal tissue. The bayonet method allows primary closure of a wound and rapid restoration of the native length of the limb

The tibial bayonet method of wound closure

Abstract Management of open lower limb fractures with soft tissue defects can be a technically challenging orthopaedic problem. Limited availability of orthoplastic services means that alternatives to the fix and flap concept are required in order to prevent infected non-unions from developing. The proposed ‘bayonet apposition’ allows the surgeon to temporarily shorten the limb without angulating the limb or creating a bone defect and removing viable bone. The viable bone edges are overlapped in a bayonet-like manner in order to appose the wound and skin edges. The limb length is restored by gradually distracting the bone segments once the soft tissues have healed. This is facilitated with a hexapod fixator for stabilization of the fracture and distraction. Prerequisites for utilizing this method are circumferential soft tissue damage to the lower limb with viable distal tissue. The bayonet method allows primary closure of a wound and rapid restoration of the native length of the limb

Monofocal distraction treatment of stiff aseptic tibial nonunions with hexapod circular external fixation

Background: Tibial nonunions remain a significant and challenging clinical problem. Nonunions occur due to a failure of biology, a failure of biomechanics, or both. The cells in and around the nonunion site have an osteogenic potential that can be activated when torsional, axial, and shearing instabilities are eliminated by establishment of an appropriate biomechanical milieu. Hexapod external fixators may be able to provide this milieu. Materials and Methods: In this retrospective study, consecutive patients with tibial nonunions treated in a single center were analyzed. They were all treated by the application of a hexapod circular fixator before deformity correction and distraction. This was followed by functional rehabilitation. We determined general health status with the use of the Short Form 12 measurement tool. Results: A total of 32 patients with 33 stiff hypertrophic nonunions were treated with circular fixators. The average age was 44 years, there were 24 males and 8 females. Out of the 33 nonunion cases, 29 united with closed distraction only; 3 required bone graft and 1 underwent amputation. The average time to union was 170 days. Overall on the final examination, all patients had a health status that was similar to the general population norm. Conclusion: A success rate in excess of 80% can be achieved with mechano-biological manipulation using hexapod external fixation in tibial nonunions without bone grafting

Functional outcomes of the failed plate fixation in distal tibial fractures salvaged by hexapod external fixator

Purpose: The purpose of this study was to evaluate the clinical and functional outcomes of failed plate fixation in distal tibia fractures salvaged with hexapod circular fixators. Materials and methods: The database of a specialized limb reconstruction center was searched for all patients with failed plate fixation undergoing limb reconstruction with a circular external fixator between 2008 and 2017. Patients between the ages of 18–65 years, with a symptomatic distal tibia malunion or non-union following plate and screw fixation were included. The SF-12 and Foot Function Index (FFI) scoring systems were used to measure clinical and functional outcomes.Results: Ten patients with a mean age of 38 ± 13 years met the inclusion criteria. Seven patients had an infected non-union, two hypertrophic non-unions, and one a malunion. The mean follow-up was 41.7 ± 28.3 months. The mean duration of external fixation was 232.9 ± 146.6 days. The SF-12 demonstrated a mean score of 49.4 ± 7.7 for the physical component and a mean score of 55.3 ± 8.1 for the mental component. Five patients (50%) scored above 45 points for the SF12 physical component, and nine patients (90%) scored above 45 points for the mental component, indicating good outcome can be achieved. The mean FFI score was 24.9 ± 19.9, and six patients had a score below 14 points (good outcome). Radiological union was observed in all 10 patients at a mean of 29 ± 14 months.Conclusions: The results of this study suggest that hexapod circular external fixation is an attractive surgical alternative for the treatment of failed plate fixation of distal tibial fractures, and can reliably achieve bony union and result in very satisfactory clinical outcomes.Level of Evidence: Level IV case series

RNAi in Arthropods: Insight into the Machinery and Applications for Understanding the Pathogen-Vector Interface

The availability of genome sequencing data in combination with knowledge of expressed genes via transcriptome and proteome data has greatly advanced our understanding of arthropod vectors of disease. Not only have we gained insight into vector biology, but also into their respective vector-pathogen interactions. By combining the strengths of postgenomic databases and reverse genetic approaches such as RNAi, the numbers of available drug and vaccine targets, as well as number of transgenes for subsequent transgenic or paratransgenic approaches, have expanded. These are now paving the way for in-field control strategies of vectors and their pathogens. Basic scientific questions, such as understanding the basic components of the vector RNAi machinery, is vital, as this allows for the transfer of basic RNAi machinery components into RNAi-deficient vectors, thereby expanding the genetic toolbox of these RNAi-deficient vectors and pathogens. In this review, we focus on the current knowledge of arthropod vector RNAi machinery and the impact of RNAi on understanding vector biology and vector-pathogen interactions for which vector genomic data is available on VectorBase

RNAi in Arthropods: Insight into the Machinery and Applications for Understanding the Pathogen-Vector Interface

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