The proteobacterial antimicrobial compound efflux (PACE) family of transport proteins was only recently described. PACE family transport proteins can confer resistance to a range of biocides used as disinfectants and antiseptics, and are encoded by many important Gram-negative human pathogens. However, we are only just beginning to appreciate the range of functions and the mechanism(s) of transport operating in these proteins. Genes encoding PACE family proteins are typically conserved in the core genomes of bacterial species rather than on recently acquired mobile genetic elements, suggesting that they confer important core functions in addition to biocide resistance. Three-dimensional structural information is not yet available for PACE family proteins. However, PACE proteins have several very highly conserved amino acid sequence motifs that are likely to be important for substrate transport. PACE proteins also display strong amino acid sequence conservation between their N— and C-terminal halves, suggesting that they evolved by duplication of an ancestral protein comprised of two transmembrane helices. In light of their drug resistance functions in Gram-negative pathogens, PACE proteins should be the subject of detailed future investigation.

Acinetobacter baumannii has rapidly emerged as a major cause of gram-negative hospital infections worldwide. A. baumannii encodes for the transport protein AceI, which confers resistance to chlorhexidine, a widely used antiseptic. AceI is also the prototype for the recently discovered proteobacterial antimicrobial compound efflux (PACE) family of transport proteins that confer resistance to a range of antibiotics and antiseptics in many gram-negative bacteria, including pathogens. The gene encoding AceI is conserved in the core genome of A. baumannii, suggesting that it has an important primordial function. This is incongruous with the sole characterized substrate of AceI, chlorhexidine, an entirely synthetic biocide produced only during the last century. Here we investigated a potential primordial function of AceI and other members of the PACE family in the transport of naturally occurring polyamines. Polyamines are abundant in living cells, where they have physiologically important functions and play multifaceted roles in bacterial infection. Gene expression studies revealed that the aceI gene is induced in A. baumannii by the short-chain diamines cadaverine and putrescine. Membrane transport experiments conducted in whole cells of A. baumannii and Escherichia coli and also in proteoliposomes showed that AceI mediates the efflux of these short-chain diamines when energized by an electrochemical gradient. Assays conducted using 8 additional diverse PACE family proteins identified 3 that also catalyze cadaverine transport. Taken together, these results demonstrate that short-chain diamines are common substrates for the PACE family of transport proteins, adding to their broad significance as a novel family of efflux pumps.

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Prevalence of zoonotic Mycobacterium bovis (bTB) disease in human population is underreported from the North of Pakistan. Here, we report on the proportion of human bTB disease among the overall TB patients, drug resistance pattern of bTB isolates, and knowledge, attitude, and practices (KAP)-based analysis of bTB disease. For this purpose, sputum samples from a total of 300 clinically diagnosed TB patients and 100 randomly selected school children suspected of pulmonary TB were processed by culture as well as polymerase chain reaction (PCR) for isolation, identification, and confirmation of Mycobacterium tuberculosis (mTB) and bTB species. Isolates of bTB were processed for drug susceptibility tests. Data on KAP regarding TB were obtained on a pretested questionnaire. Sputum-based PCR results indicated that 288/300 (96%) were confirmed as mTB, while 12/300 (4%) were found as bTB diseases. Interestingly, none of the school child was declared positive for either mTB or bTB. Notably, 274/300 (91.3%) positively cultured samples were identified as mTB, 13/300 (4.3%) as bTB, while 5/300 (1.7%) as mixed containing both. Importantly, except one, all of the bTB isolates were found resistant to pyrazinamide. Surprisingly, most of the bTB isolates (~70%) were found resistant to a broad range of first- and second-line anti-TB drugs. SplitsTree and recombination analysis indicated no evidence of intergenic recombination. Finally, residence, occupation, presence of animals at home, and sleeping alongside animals were found significantly associated with occurrence of bTB disease. To the best of our knowledge, we report for the first time on the high (4%) burden of bTB disease in human TB patients in Peshawar, Pakistan.

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The objective of this study was to conduct a qualitative analysis of raw beef meat sold in the city of Quetta, Pakistan for presence and drug sensitivity of the potentially pathogenic Escherichia coli strain O157:H7. The study used 200 raw beef meat samples collected from retail butcher shops. Conventional and rapid biochemical tests, latex agglutination and multiplex polymerase chain reaction (PCR) using primers designed for the rfb O157 and flic H7 genes were used to detect E. coli O157:H7. All O157:H7 isolates were also tested for Shiga toxin genes stx 1 and stx 2. The prevalence of E. coli O157:H7 in collected beef meat samples was 10%. Detection through PCR was found more sensitive than detection of O and H antigens. The quantity of E. coli O157:H7 isolates positive for Shiga toxins was 50% (20% for stx 1, 45% for stx 2 and 10% for both stx 1 and stx 2). Season wise variation showed highest E. coli O157:H7 prevalence during summer months. A further concern is that E. coli O157:H7 isolates were resistant to a range of common antibiotics. The results indicate an urgent need for applying proper food hygiene practices in the Quetta region to reduce incidence of foodborne diseases and they also emphasize the global problem of antimicrobial resistance. Practical applications E. coli O157:H7 is as a potentially threatening foodborne pathogen. A significant prevalence of E. coli O157:H7 detected in raw beef meat from retail outlets in the city of Quetta indicates an urgent need for applying proper food hygiene practices in the Quetta region to reduce the incidence of foodborne diseases. Furthermore, resistance of the E. coli O157:H7 isolates to a range of commonly used antibiotics emphasizes the global problem of antimicrobial resistance. The multiplex PCR method used here is a reliable, sensitive, and relatively rapid technique for detecting E. coli O157:H7 in food and environmental samples and important for ongoing surveillance to minimize contamination of raw meat products and associated cross contamination by E. coli O157:H7.

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