Prognostic significance of novel katG mutations in Mycobacterium tuberculosis.

BACKGROUND: By using whole genome sequencing (WGS), researchers are beginning to understand the genetic diversity of Mycobacterium tuberculosis (MTB) and its consequences for the diagnosis of multidrug-resistant tuberculosis (MDR-TB) on a genomic scale. The Global Consortium for Drug-resistant TB Diagnostics (GCDD) conducted a genome scale variant analyses of 366 clinical MTB genomes (mostly MDR/XDR [extensively drug resistant]) from four countries in order to inform the development of rapid molecular diagnostics. This project has been extended by performing an evolutionary analysis of isoniazid (INH)-resistant isolates for prognostic purposes. METHODS: 151 (130 INHR, 21 INHS) clinical MTB isolates from India (19: 17 INHR, 2 INHS), Moldova (48: 42 INHR, 6 INHS), the Philippines (26: 20 INHR, 6 INHS), and South Africa (58: 51 INHR, 7 INHS) were included in this study. INH drug susceptibility was determined by using MGIT 960 and WHO (World Health Organization)-recommended critical concentration of 0.1 mg/L. Isolates were sequenced using PacBio RS WGS platform. A genome-wide variant analysis was conducted using a proprietary pipeline (PacDAP) developed at San Diego State University. To infer the amino acid changes in katG that confer resistance, PAML was utilized to detect sites in silico that are under positive selection. The dN/dS method was used in combination with Bayes empirical Bayes to determine sites under positive selection and Chi-Squared analysis to determine the significance of the selected sites. RESULTS: PacDAP variant analysis revealed 22 novel catalase-peroxidase (katG product) mutations. Of these, 14 were single nucleotide polymorphisms, while 8 novel mutations appeared in combination with katG S315T and/or with inhA promoter C-15T. These SNPs have not been previously reported. Additionally, 11 previously observed, but uncommon, katG mutations were also observed in these clinical isolates. These results suggest that 17 amino acids in the enzyme are under positive selective pressure; most significantly in South Africa and the Philippines. No selective pressure on codons other than 315 was observed in isolates from Moldova. Due to the low number of isolates from India, the significance of the sites under positive selection was low and no prediction for India could be made based on this study. CONCLUSIONS: Eleven of the 14 SNPs are resistance conferring, and it is believed that the remaining 8 combinatorial mutations are either compensatory in nature or, in combination with known SNPs, could increase resistance levels. Positive selection results indicate a diversifying evolutionary path to resistance more in line with long tail statistics and therefore indicate a departure from the traditional point mutation (or "hotspot") model that current molecular diagnostics are based on. Positive selection pressures indicate a future with elevated diagnostic and prognostic significance of the "long tail" (i.e., alternative mechanisms of resistance) and potentially diminishing significance of the canonical mutations (especially in South Africa and the Philippines), which could have significant future implications on narrowly targeting molecular diagnostics.

Veterinary practice internal controls.

This chapter explores the responsive management requirements critical for effective internal controls. Internal control techniques offered include the application of segregation of function, goal-orientation, performance planning, and training objectives in five veterinary practice areas. These areas are client controls, cash controls, inventory controls, purchase controls, and stock controls. Because only trusted staff members are given access to practice assets, the veterinary leadership must have a responsive management plan to protect the life of the practice while building the practice team. Methods are shared for controlling opportunities that can lead to employee fraud while concurrently building a team approach to internal controls.

A survey on the question of how well veterinarians are prepared to predict their client's human-animal bond.

In a survey of 63 military veterinary facilities, actual responses from 961 clients were compared with 86 servicing US Army Veterinary Corps practitioners' predictions of the clients' responses. The survey questions were designed to evaluate the degree of human-animal bonding of the military clients. The survey revealed that the effects of human-animal interaction on community, family, and animal health have extensive impact on the military member, the military installation, and the military health care delivery system. Results revealed sociologic and psychologic factors that influenced the quality of life of mobile military members and their families. The survey also revealed that although the newly graduated veterinarians understood the professional and scientific values of their clients, they often underestimated the social interdependence between client and pet. The military veterinarians in this survey were considered to be representative of the new graduate, and the results are relevant to any civilian practitioner concerned with client relations or hiring new graduates.