Right foot after 14 days of treatment with amikacin and trimethoprim-sulfamethoxazole.

<p>Sinuses were closed and no more discharge observed.</p

Pictures A, B and C show the right foot before treatment, Pictures D, E and F show the foot 22 weeks after treatment.

<p>Pictures A, B and C show the right foot before treatment, Pictures D, E and F show the foot 22 weeks after treatment.</p

Western blot performed from the first patient's serum before and after cross-adsorption.

<p>WM = Weight marker; lanes 1, 3 and 5: <i>B. quintana</i> antigen; lanes 2, 4 and 6: <i>B. henselae</i> antigen; lanes 1 and 2: unadsorbed serum; lanes 3 and 4: serum adsorbed with <i>B. henselae</i>; lanes 5 and 6: serum adsorbed with <i>B. quintana</i>.</p

Overview of confirmed <i>N</i>. <i>sennetsu</i> patients in Laos.

<p>Overview of confirmed <i>N</i>. <i>sennetsu</i> patients in Laos.</p

<i>Neorickettsia sennetsu</i> as a Neglected Cause of Fever in South-East Asia

<div><p><i>Neorickettsia sennetsu</i> infection is rarely recognized, with less than 100 globally reported patients over the last 50 years. The disease is thought to be contracted by eating raw fish, a staple of many South-East Asian cuisines. In 2009, the first patient with sennetsu was identified in the Lao PDR (Laos), raising the question as to how common this organism and related species are in patients presenting with fever. We investigated the frequency of <i>N</i>. <i>sennetsu</i> infection at hospitals in diverse areas of Laos. Consenting febrile hospital inpatients from central (Vientiane: n = 1,013), northern (Luang Namtha: n = 453) and southern (Salavan: n = 171) Laos were screened by PCR for <i>N</i>. <i>sennetsu</i>, if no previous positive direct diagnostic test was available. A PCR-restriction fragment length polymorphism assay was developed to differentiate between <i>N</i>. <i>sennetsu</i>, <i>Ehrlichia chaffeensis</i> and <i>Anaplasma phagocytophilum</i>. To allow more detailed studies of <i>N. sennetsu</i>, culture was successfully established using a reference strain (ATCC VR-367), identifying a canine-macrophage cell line (DH82) to be most suitable to visually identify infection. After screening, <i>N</i>. <i>sennetsu</i> was identified and sequence confirmed in four (4/1,637; 0.2%) Lao patients. Despite the previously identified high seroprevalence of <i>N</i>. <i>sennetsu</i> antibodies in the Lao population (~17%), acute <i>N</i>. <i>sennetsu</i> infection with sufficient clinical signs to prompt hospitalization appears to be rare. The reservoir, zoonotic cycle and pathogenicity of <i>N</i>. <i>sennetsu</i> remain unclear and require further investigations.</p></div

Electron microscopic appearance of <i>N</i>. <i>sennetsu</i> (ATCC VR-367, Miyayama strain) in DH82 canine monocyte cultures. a)

<p>Low power micrograph of an infected cell in which a number of bacteria can be identified in the cytoplasm (arrowheads) in addition to the nucleus (N), mitochondria (Mi) and lipid droplet (L). Note the single extracellular bacterium (arrow). Bar represents 1μm. <b>b)</b> Enlargement of part of the cytoplasm showing a number of gram negative bacteria (B). N–nucleus. Bar represents 200nm. <b>c)</b> Detail of N. sennetsu (arrow) showing the gram negative bacteria limited by two unit membranes located within a membrane bound vacuole. G—Golgi stack. Bar represents 200nm.</p

Schematic alignment of <i>N</i>. <i>sennetsu</i> and related organisms showing the restrictions sites used for the RFLP.

<p>The unique RFLP pattern of the 16 sRNA gene target, after incubation with <i>Alu</i>I (green), <i>Bsm</i>FI (yellow) or <i>Sty</i>I (red) allow the differentiation of <i>N</i>. <i>sennetsu</i>, <i>E</i>. <i>chaffeensis</i> and <i>A</i>. <i>phagocytophium</i> as well as other potentially amplified organisms. The resulting fragment sizes are as follows; <i>N</i>. <i>sennetsu</i>–<i>Alu</i>I: 345bp (uncut); <i>Bsm</i>F1: 180bp, 80bp, 60bp, 16bp; <i>Sty</i>I: 215bp, 127bp; <i>E</i>. <i>chaffeensis–Alu</i>I: 345bp (uncut); <i>Bsm</i>F1: 328bp, 16bp; <i>Sty</i>I: 215bp, 127bp; <i>A</i>. <i>phagocytophium–Alu</i>I: 199bp, 145bp <i>Bsm</i>F1: 328bp, 16bp; <i>Sty</i>I: 345bp (uncut).</p