Mycobacterium kansasii has emerged as an important nontuberculous mycobacterium pathogen, whose incidence and prevalence have been increasing in the last decade. M. kansasii can cause pulmonary tuberculosis clinically and radiographically indistinguishable from that caused by Mycobacterium tuberculosis infection. Unlike the widely-studied M. tuberculosis, little is known about the innate immune response against M. kansasii infection. Although inflammasome activation plays an important role in host defense against bacterial infection, its role against atypical mycobacteria remains poorly understood. In this report, the role of inflammasome activity in THP-1 macrophages against M. kansasii infection was studied. Results indicated that viable, but not heat-killed, M. kansasii induced caspase-1-dependent IL-1β secretion in macrophages. The underlying mechanism was found to be through activation of an inflammasome containing the NLR (Nod-like receptor) family member NLRP3 and the adaptor protein ASC (apoptosis-associated speck-like protein containing a CARD). Further, potassium efflux, lysosomal acidification, ROS production and cathepsin B release played a role in M. kansasii-induced inflammasome activation. Finally, the secreted IL-1β derived from caspase-1 activation was shown to restrict intracellular M. kansasii. These findings demonstrate a biological role for the NLRP3 inflammasome in host defense against M. kansasii.

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Introduction: Laryngeal swab samples collected from three waterfowl slaughterhouses in central Taiwan were cultured and suspected isolates of Riemerella anatipestifer were identified by API 20NE and 16S rDNA PCR. Material and Methods: Serum agglutination was used for serotyping, and antimicrobial susceptibility was tested. Results: Seventy-six R. anatipestifer isolates were detected, and the prevalences in the ducks and geese were 12.3% (46/375) and 8.0% (30/375), respectively. The positive isolation rates were 65.6% for all arriving waterfowl, 76.0% for birds in the holding area, 1.6% for defeathered carcasses, but zero for degummed carcasses. A PCR examination detected R. anatipestifer in the slaughtering area frequently. Serotype B was dominant in both duck (34.8%) and goose (46.7%) isolates, but the wide serotype distribution may very well impede vaccination development. All isolates were resistant to colistin, and 79.7% were resistant to more than three common antibiotics. Conclusion: The results proved that most ducks had encountered antibiotic-resistant R. anatipestifer in rearing, which suggests that the bacterium circulates in asymptomatic waterfowl. It is worth noting that most waterfowl farms were found to harbour R. anatipestifer, and contaminated slaughterhouses are a major risk factor in its spread. Effective prevention and containment measures should be established there to interrupt the transmission chain of R. anatipestifer.

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IS629 is 1,310 bp in length with a pair of 25-bp imperfect inverted repeats at its termini. Two partially overlapping open reading frames, orfA and orfB, are present in IS629, and two putative translational frameshift signals, TTTTG (T 4 G) and AAAAT (A 4 T), are located near the 3-end of orfA. With the lacZ gene as the reporter, both T 4 G and A 4 T motifs are determined to be a ؊1 frameshift signal. Two peptides representing the two transframe products designated OrfAB and OrfAB, are identified by a liquid chromatography-tandem mass spectrometric approach. Results of transposition assays show that OrfAB is the transposase and that OrfAB aids in the transposition of IS629. Pulse-chase experiments and Escherichia coli twohybrid assays demonstrate that OrfAB binds to and stabilizes OrfAB, thus increasing the transposition activity of IS629. This is the first transposable element in the IS3 family shown to have two functional frameshifted products involved in transposition and to use a transframe product to regulate transposition.The insertion sequence IS629 is a member of the IS3 family of transposable elements. It was initially isolated from the chromosome of Shigella sonnei (1) and has been detected in many other enteric bacteria, including S. dysenteriae, S. flexneri, S. boydii, Escherichia coli C, E. coli O157:H7, Enterobacter cloacae MD36, and Serratia marcescens (2). IS629 is 1,310 bp in length and has a pair of 25-bp imperfect inverted repeats at its termini (3). Similar to the genetic organization of other members of the IS3 family, two consecutive and partially overlapping open reading frames, designated orfA and orf B, are present in IS629 (see Fig. 1). The coding potential of orfA (nucleotides 55-381) is 108 amino acids, and that of orf B (nucleotides 378 -1,268) is 296 amino acids (3). The stop codon ( 379 TGA) of orfA overlaps the initiation codon ( 378 ATG) of orf B (see Fig. 1). A putative promoter and the Shine-Dalgarno sequence are found upstream from the initiation codon of orfA, but no such sequences are present in the upstream region of orf B (3).Two putative Ϫ1 translational frameshift signals, TTTTG (T 4 G) and AAAAT (A 4 T), are located near the 3Ј-end of the orfA at nucleotide positions 342-346 and 375-379, respectively ( Fig. 1) (4), suggesting the existence of two frameshifted products. In this study, we demonstrated that both of these two putative frameshift signals are functional, causing a Ϫ1 translational frameshift and resulting in the production of two transframe products designated OrfABЈ and OrfAB. OrfABЈ was shown to be the transposase of IS629, and OrfAB was demonstrated to bind and stabilize OrfABЈ. EXPERIMENTAL PROCEDURESCloning of IS629-A fragment containing both orfA and orf B sequences of IS629 was amplified by PCR from the chromosome of S. sonnei (ATCC 9290) or E. coli O157 with primers F NdeI-55 and R Ecl136II-Term (Table 1). The PCR product thus generated was cloned into pGEMT-Easy (Promega) to produce pGEMT629. Subsequently, the 1.2-kb NdeI-Ecl136II fragment containing...