Acid-fast bacterium detection and identification from paraffin-embedded tissues using a PCR-pyrosequencing method.

AIMS: Acid-fast bacterium (AFB) identification from formalin-fixed paraffin-embedded (FFPE) tissues is challenging and may not be readily available to the clinical laboratory. A method to detect and identify AFB from FFPE tissues using PCR and pyrosequencing (PCR-Seq) was developed and evaluated. METHODS: The method was validated using spiked cell-clotted paraffin blocks before use with patients' specimens. DNA was extracted from tissue sections, and a 16S rRNA gene fragment was amplified and a signature sequence was produced on a PyroMark ID system. Sequences were aligned to established databases for AFB identification. Additional tissue sections were stained and examined for AFB. RESULTS: Both sensitivity and specificity were 100% on spiked cell-clotted blocks without cross-reactivity with non-AFB. Of 302 FFPE tissues from patients, 116 (38%) were AFB-stain positive; 83 (72%) of these had AFB identified. The 21 AFB identified included Mycobacterium tuberculosis complex (14 cases), Mycobacterium leprae (3), Mycobacterium genavense (2), Mycobacterium marinum-ulcerans group (3) and 17 other AFB (61). Thirteen cases were AFB-stain indeterminate and 4 were positive by the PCR-Seq method. Of the AFB stain-negative cases, 167 were negative and 6 were positive by PCR-Seq. CONCLUSIONS: The PCR-Seq method provided specific identification of various AFB species or complexes from FFPE tissues.

Influence of in-house rehabilitation on the postoperative outcome of dogs with intervertebral disk herniation.

OBJECTIVE: To determine the influence of in-house rehabilitation on the postoperative outcome of dogs with intervertebral disk herniation (IVDH). DESIGN: Retrospective cohort study. ANIMALS: Dogs (n = 248) under 20 kg of weight, with single site, thoracolumbar, Hansen Type I IVDH. PROCEDURES: Medical records of non-ambulatory dogs with IVDH treated via hemilaminectomy were reviewed. Dogs were classified in 2 groups depending on whether their postoperative management included an in-house rehabilitation program. Preoperative and sequential postoperative modified Frankel scores (MFSs) were recorded. Time to ambulation, time to normal conscious proprioception, final MFS, and complications were compared between the groups. RESULTS: More dogs returned to full neurologic function (final MFS of Grade 5) when in-house rehabilitation was included in the postoperative management (33% compared to 9%). Normal conscious proprioception and ambulation returned earlier in the control group (42 days and 14 days, respectively) compared to the group with rehabilitation (49 days and 28 days, respectively). The complication rate was higher in the control group (29%) compared with the group with rehabilitation (16%). CONCLUSIONS AND CLINICAL RELEVANCE: In-house rehabilitation should be included in the postoperative management in dogs after surgical treatment of IVDH to improve neurologic function and reduce postoperative complications.

Location of the extracranial extent of leprous facial nerve pathology may allow leprous facial palsy to be reanimated by free muscle transfer.

Leprosy is a mycobacterial nerve and skin infection, which can be eradicated by antibiotics. Some patients affected by leprosy, once cured, have residual nerve impairment with paralysis and sensory neuropathy. A series of patients with facial nerve paralysis, investigated using clinical, histological and electrophysiological techniques, demonstrated that the nerve pathology was distal to the section of main trunk prior to its bifurcation. Facial reanimation was achieved with a free gracilis-muscle transfer, coapting its motor nerve to the ipsilateral facial nerve trunk proximal to the site of the leprosy pathology, with a moderate clinical result.

Location of the extracranial extent of leprous facial nerve pathology may allow leprous facial palsy to be reanimated by free muscle transfer

Leprosy is a mycobacterial nerve and skin infection, which can be eradicated by antibiotics. Some patients affected by leprosy, once cured, have residual nerve impairment with paralysis and sensory neuropathy. A series of patients with facial nerve paralysis, investigated using clinical, histological and electrophysiological techniques, demonstrated that the nerve pathology was distal to the section of main trunk prior to its bifurcation. Facial reanimation was achieved with a free gracilis-muscle transfer, coapting its motor nerve to the ipsilateral facial nerve trunk proximal to the site of the leprosy pathology, with a moderate clinical result.

Leprosy affects the tibial nerves diffusely from the middle of the thigh to the sole of the foot, including skip lesions.

This study investigated where leprosy affects the posterior tibial nerve and whether neurolysis is beneficial. Nine patients with bilateral posterior tibial leprous neuropathy with no sensorimotor recovery were studied. Preoperative sensory-muscle and nerve conduction velocity testing revealed the tarsal tunnel to be the site of a severe lesion in all cases. During surgery, the most proximal site of the nerve lesion was detected by electrically stimulating the spinal roots from the second lumbar nerve to the fourth sacral nerve, evoking efferent mixed nerve compound action potentials that were recorded from the exposed tibial nerve. In all patients, the nerve compound action potentials became normal only proximal to the sciatic nerve bifurcation. Epineuriotomy within these seemingly unaffected segments revealed fibrosis of the interfascicular epineurium. Interfascicular neurolysis was performed on all affected segments. A 2-year follow-up showed an increase in girth of the proximal calf musculature in six of eight patients (the ninth patient had no recordable nerve conduction velocity). It was concluded that (1) leprosy affects the tibial nerves in a scattered way from the sciatic nerve main trunk distally to the exit of the tarsal tunnel; and (2) interfascicular, microsurgical neurolysis is beneficial provided that it is performed on all affected nerve segments.

The stapedial reflex as a topographical marker of proximal involvement of the facial nerve in leprosy. A pilot study.

This study aimed to determine the parameters necessary for a study of stapedial reflexes in leprosy patients to ascertain if the facial nerve is involved more proximally than the stylomastoid foramen. It involved leprosy patients with and without facial nerve involvement and non-leprosy controls. Clinical examination of the patients' ears, a tympanogram and audiogram to exclude conductive and sensorineural deafness, followed by the measurement of a stapedial reflex and the acoustic reflex threshold, were carried out. The number of absent reflexes and the acoustic reflex thresholds did not differ between the three groups of subjects. A definitive study would be logistically impossible. Suggestions are made as to more exact patient selection in order to demonstrate any stapedial reflex changes due to leprosy. The findings of this study do not suggest that facial nerve pathology extends proximally to the stylomastoid foramen, unless such proximal involvement is subclinical to the detection methods used.

Facial nerve pathology in leprosy: searching for the proximal extent of the lesion in facial nerve biopsies.

A light and electron microscope study was made of resin embedded facial nerves in three cases of leprosy involving the facial nerve. The patients had irreversible facial nerve palsies and had requested facial reconstruction. No consistent pattern of nerve fibre damage was found. In one case the temporozygomatic was affected, but the cervical branch was normal, suggesting the damage begins distally. In two cases the loss of nerve fibres in the trunk and all branches was similar, and is likely to emanate from damage at a more proximal site. The presence of increased numbers of unmyelinated axons, often in clusters, is evidence of regeneration. These axons probably have the potential to develop into functional myelinated fibres provided that they can innervate a viable distal target such as a muscle graft. These regenerating axons are distal to the stylomastoid foramen suggesting that the most proximal level of involvement of the facial nerve could be intracranial. The finding of a more proximal level of nerve involvement, implies that the mis-reinnervation seen in partially recovered facial nerve palsies in leprosy, could be due to some regenerating axons being mis-directed at the level of the main trunk bifurcation.

Cervical branch of the facial nerve in leprosy.

This study demonstrates that the platysma is occasionally palsied in leprosy and that this only occurs when the facial nerve already has some other palsy. That there needs to be a facial palsy before there can be a platysma palsy is strongly suggested, in that there was no case of an isolated platysma palsy. Patients, regardless of age or other factors, could mimic a platysma contraction. This obviates the need for electrical testing to examine for a platysma palsy. It also means that a nonfunctioning platysma on clinical examination is, in fact, a palsied platysma. While lagophthalmos is regularly examined for, and any obvious facial paresis would be noticed, less severe forms of facial muscle paresis will only be found if formally examined for. The mechanism whereby the facial nerve is involved in leprosy is not clarified, but our findings suggest that proximal spread of a lesion that began in the zygomatico-temporal branches and reaches to the facial nerve trunk is more likely than new lesions developing de novo in other peripheral facial nerve branches. That the primary lesion is within the facial nerve trunk in all cases but we only see the frequent zygomatic sequelae due to secondary factors is not excluded.

The distally based posterior interosseous fasciocutaneous island flap in reconstruction of the hand in leprosy.

In leprosy a functionally useless hand can be the sequelae of resorption of the sensation-impaired thumb and fingers resulting from repeated trauma and infection. When this process shortens the thumb to the level of the proximal phalanx or metacarpal, the effect is to produce a relatively shallow first web space which together with the shortening, can prevent the most basic hand manoeuvre of a pincer or pinch grip. The reconstructive procedure commonly used in this situation is to widen and deepen the web space with a z-plasty combined with excision of the second metacarpal but the result can be inadequate. We have used the posterior interosseous fasciocutaneous island flap, both as a simple and a compound flap, to solve this challenging problem and we report here our experience with four patients.

Interosseous transfer of tibialis posterior for common peroneal nerve palsy.

The interosseous route remains popular for tibialis posterior tendon transfer for drop-foot. It leaves a smaller range of movement than the circumtibial route, but lengthening the calcaneal tendon may improve this. The results of this present series indicate that, in order to predict a good functional result, the ankle must be held in at least 20 degrees of dorsiflexion at the time of tendon transfer.