The susceptibility of Leptospira interrogans serovar icterohaemorrhagiae strain Verdun to selected antibiotics used in medical practice (ampicillin, doxycycline, and ofloxacin) was evaluated in a Syrian hamster model, to determine the efficacy of these antibiotics during the course of the disease. A quantitative PCR assay was used to monitor the density of leptospires in blood and in target organs (liver, kidney, lung, heart, and spleen). Our results demonstrated the ability of ampicillin at a high dose (100 mg/kg of body weight) to clear leptospires from the host, except from kidneys and heart, where 10 2 leptospires/g remained at day 6. Ofloxacin (30 mg/kg) was unable to clear bacteria from blood or kidneys. With doxycycline (10 mg/kg), the clearance of leptospires occurred in 2 days in all the target organs studied, with the exception of liver, which required 3 days. Our data demonstrate the value of monitoring the kinetics of experimental leptospiral infection in order to accurately evaluate the efficacy of antibiotics. We have demonstrated the potential value of doxycycline for the treatment of leptospirosis cases, except in circumstances where it is contraindicated. This experimental model could be used to define better therapeutic strategies for human leptospirosis, by testing associations or new formulations of antibiotics.

The Vanuatu archipelago is endemic for HTLV-1 with a diversity of subtype C variants. These strains were probably introduced into Vanuatu during ancient migration of the original settlers a few thousand years ago.

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BackgroundHuman T-Lymphotropic Virus type 1 (HTLV-1) is endemic among people of Melanesian descent in Papua New Guinea, Solomon Islands and Vanuatu, and in Indigenous populations from Central Australia. Molecular studies revealed that these Australo-Melanesian strains constitute the highly divergent HTLV-1c subtype. New Caledonia is a French overseas territory located in the Southwest Pacific Ocean. HTLV-1 situation is poorly documented in New Caledonia and the molecular epidemiology of HTLV-1 infection remains unknown.ObjectivesStudying 500 older adults Melanesian natives from New Caledonia, we aim to evaluate the HTLV-1 seroprevalence and to molecularly characterize HTLV-1 proviral strains.Study designPlasma from 262 men and 238 females (age range: 60–96 years old, mean age: 70.5) were screened for anti-HTLV-1 antibodies by particle agglutination (PA) and indirect immunofluorescence assay (IFA). Serological confirmation was obtained using Western blot assay. DNAs were extracted from peripheral blood buffy coat of HTLV-1 seropositive individuals, and subjected to four series of PCR (LTR-gag; pro-pol; pol-env and tax-LTR). Primers were designed from highly common conserved regions of the major HTLV-1 subtypes to characterize the entire HTLV-1 proviral genome.ResultsAmong 500 samples, 3 were PA and IFA positive. The overall seroprevalence was 0.6%. The DNA sample from 1 New Caledonian woman (NCP201) was found positive by PCR and the complete HTLV-1 proviral genome (9,033-bp) was obtained. The full-length HTLV-1 genomic sequence from a native woman from Vanuatu (EM5), obtained in the frame of our previous studies, was also characterized. Both sequences belonged to the HTLV-1c Australo-Melanesian subtype. The NCP201 strain exhibited 0.3% nucleotide divergence with the EM5 strain from Vanuatu. Furthermore, divergence reached 1.1% to 2.9% with the Solomon and Australian sequences respectively. Phylogenetic analyses on a 522-bp-long fragment of the gp21-env gene showed the existence of two major clades. The first is composed of strains from Papua New Guinea; the second includes strains from all neighboring archipelagos (Solomon, Vanuatu, New Caledonia), and Australia. Interestingly, this second clade itself is divided into two sub-clades: strains from Australia on one hand, and strains from Solomon Islands, Vanuatu and New Caledonia on the other hand.ConclusionsThe HTLV-1 seroprevalence (0.6%) in the studied adult population from New Caledonia appears to be low. This seroprevalence is quite similar to the situation observed in Vanuatu and Solomon Islands. However it is very different to the one encountered in Central Australia. Taken together, these results demonstrated that Australo-Melanesia is endemic for HTLV-1 infection with a high diversity of HTLV-1c strains and a clear geographic clustering according to the island of origin of HTLV-1 infected persons.

Fluoroquinolone resistance in Streptococcus pneumoniae mainly involves stepwise mutations predominantly in the parC and gyrA genes. We have developed a single-run real-time PCR assay for detection of the four most common mutations in the quinolone resistance-determining regions of these genes. This assay provides a useful tool for both clinical and epidemiological use.Streptococcus pneumoniae is a major cause of communityacquired infections ranging from otitis media to pneumonia or meningitis (12). Strains resistant to multiple antibiotics have recently been reported and pose a risk of therapeutic failure (1). This observation has encouraged the use of newer fluoroquinolones against pneumococci (8) and has thus led to the emergence of fluoroquinolone-resistant clones. Prevalence rates have increased in Canada and have reached 14.3% in Hong Kong, while they have remained below 1 to 2% in the United States and Europe (3). The development of resistance to fluoroquinolones in pneumococci is a stepwise mutational process (7) that mainly occurs in the quinolone resistancedetermining regions (QRDRs) of the parC and gyrA genes (11). Several reports have noted that a significant proportion of isolates harboring first-step mutations had low or no phenotypic expression but had the potential to develop higher levels of resistance to fluoroquinolones when they suffered a second mutation, resulting in treatment failure (10). This evolution implies that microbiologists must be able to detect first-step, or "preresistant," mutants. The use of a nonmolecular test scheme for the detection of low-level resistance has been proposed, but this type of test lacks sensitivity and specificity (13). A powerful and highly specific technique for the detection of mutations within a DNA sequence is real-time PCR with fluorescent resonance energy transfer (FRET) hybridization probes. We report on the development of a real-time PCR assay that uses FRET probes for the single-run detection of the four most common resistance mutations within the QRDRs of the parC and gyrA genes of S. pneumoniae.The S. pneumoniae strains were isolated, cultured, and identified as described previously (9). A total of 58 strains whose parC and gyrA genes had been sequenced previously were studied. They consisted of strain R6 and 3 R6 derivatives kindly provided by E. Varon (13), 10 mutants generated in vitro, reference strains CIP104485 and CP 1000, and 42 clinical isolates (37 from New Caledonia, 3 from Australia, and 2 from Tahiti). Table 1 summarizes the known mutations, the results of disk agar diffusion (Bio-Rad), and the MICs determined by agar dilution, as described previously (13). DNA was extracted with a QIAamp DNA mini kit (Qiagen). The primers and probes were designed by using LightCycler probe design software (version 2) and were ordered from Proligo Singapore Pty. Ltd. The wild-type sequences of gyrA (GenBank accession number DQ175176) and parC (GenBank accession number DQ176507) were used for oligonucleotide design. By using the parC sequence, an LC-R...