Concurrent chemoradiation for locally advanced stage III non-small cell lung cancer with cisplatin, vinorelbine, and thoracic radiotherapy: a phase II study from the Galician Lung Cancer Group

Purpose: The aim of this phase II study was to evaluate the activity and safety of the combination of cisplatin and vinorelbine with thoracic radiotherapy in unresectable locally advanced stage III non-small cell lung cancer (NSCLC). The primary endpoint was the objective response rate (ORR). Secondary objectives included toxicity profile, progression-free survival (PFS), and overall survival (OS). Materials and methods: A total of 48 NSCLC patients were enrolled (median age 60 years, 52% stage IIIA and 48% stage IIIB, 52% adenocarcinoma). Patients received three cycles of chemotherapy every 21 days [intravenous cisplatin 80 mg/m2 and intravenous vinorelbine 25 mg/m2 on day 1 and oral vinorelbine on day 8 (60 mg/m2)] concurrent with radiotherapy (66 Gy, administered at 1.8 Gy per day, five consecutive days per week). Results: ORR was 79.2% (72.9% showing partial response and 6.3% showing complete response). With a median follow-up of 20.7 months, median PFS was 12 months and median OS was 36 months. Grade 3/4 toxicities were: neutropenia (14.5%), anaemia (6.2%), vomiting (2%), and oesophagitis (4.2%). No toxic deaths were reported. Conclusion: This combined regimen shows efficacy and a manageable safety profile. PFS and OS outcomes are encouraging and warrant further research

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Concurrent chemoradiation for locally advanced stage III non-small cell lung cancer with cisplatin, vinorelbine, and thoracic radiotherapy: a phase II study from the Galician Lung Cancer Group.

PURPOSE: The aim of this phase II study was to evaluate the activity and safety of the combination of cisplatin and vinorelbine with thoracic radiotherapy in unresectable locally advanced stage III non-small cell lung cancer (NSCLC). The primary endpoint was the objective response rate (ORR). Secondary objectives included toxicity profile, progression-free survival (PFS), and overall survival (OS). MATERIALS AND METHODS: A total of 48 NSCLC patients were enrolled (median age 60 years, 52% stage IIIA and 48% stage IIIB, 52% adenocarcinoma). Patients received three cycles of chemotherapy every 21 days [intravenous cisplatin 80 mg/m2 and intravenous vinorelbine 25 mg/m2 on day 1 and oral vinorelbine on day 8 (60 mg/m2)] concurrent with radiotherapy (66 Gy, administered at 1.8 Gy per day, five consecutive days per week). RESULTS: ORR was 79.2% (72.9% showing partial response and 6.3% showing complete response). With a median follow-up of 20.7 months, median PFS was 12 months and median OS was 36 months. Grade 3/4 toxicities were: neutropenia (14.5%), anaemia (6.2%), vomiting (2%), and oesophagitis (4.2%). No toxic deaths were reported. CONCLUSION: This combined regimen shows efficacy and a manageable safety profile. PFS and OS outcomes are encouraging and warrant further research.

Isolation of Naegleria fowleri from a domestic water tank associated with a fatal encephalitis in a 4 month-old Venezuelan child.

This study describes the association of household water system contamination with the pathogenic Free-Living Amoeba (FLA) Naegleria fowleri and a case of fatal Primary Amoebic Meningoencephalitis (PAM) in a child from the state of Monagas in Venezuela. Amoebae were initially identified by microscopy from a sample of cerebrospinal fluid (CSF) from the child. Direct DNA extraction and specific PCR/sequencing for N. fowleri was also carried out from the same CSF sample. In order to determine a possible environmental source of infection, water samples from the water tank of the child's home and also water bodies recently visited by the child and his family, were examined for the presence of N. fowleri by culture and PCR/sequencing. The results obtained from the collected water samples revealed that only the water tank of the house was positive for N. fowleri. PCR/sequencing showed that the strains isolated from the patient and the water tanks were 100 % identical. Therefore, the house water tank was confirmed as the source of infection in this case, possibly as a result of the occasional immersion of the child´s head under the water while bathing. This case highlights a novel source of thermally polluted water and another threat of N. fowleri infection.

Balamuthia mandrillaris therapeutic mud bath in Jamaica.

Balamuthia mandrillaris is an emerging cause of encephalitis in humans. The transmission dynamics are poorly understood due to the high fatality rate and the sporadic nature of cases. Seventy-two soil samples were collected from beaches and the banks of lagoons, rivers, ponds, mineral springs and streams from across Jamaica and assayed for the presence of B. mandrillaris. Seventy-nine sites were sampled and the mitochondrial 16S rDNA gene of B. mandrillaris was amplified and sequenced to confirm the presence of the amoeba. One isolate of B. mandrillaris was recovered from soil from mineral spring which hosts an informal therapeutic mud bath business. Although B. mandrillaris is less frequently isolated from soil than other free-living amoebae, rubbing mud containing the organism onto the skin increases the likelihood of exposure and infection. This first report on the isolation of B. mandrillaris in the Caribbean and its presence in soil where human contact is likely warrants further investigation using serological methods to elucidate exposure patterns.

Effect of postoperative physical training on activity after curative surgery for non-small cell lung cancer: a multicentre randomised controlled trial.

OBJECTIVE: To evaluate the effect of a combined hospital plus home exercise programme following curative surgery for non-small cell lung cancer (NSCLC). DESIGN: Randomised controlled trial. SETTING: Teaching hospital. PARTICIPANTS: One hundred and thirty-one subjects with NSCLC admitted for curative surgery. INTERVENTIONS: Participants were randomised to usual care or a hospital plus home exercise programme. OUTCOMES: The primary outcome was the between-group difference in physical activity 4 weeks after surgery. Secondary outcomes were the difference in quadriceps strength, exercise tolerance and quality of life [Short Form-36 (SF-36) and European Organisation for Research and Treatment of Cancer (EORTC) QLQ-LC13] from pre-operatively (baseline) to 4 weeks after surgery. RESULTS: The participants (n=131) had a mean age of 68 [standard deviation (SD) 11] years and mean forced expiratory volume in 1 second of 2.4 (SD 1.1)l. There were no significant differences in physical activity between the groups 4 weeks after surgery [mean difference adjusted for baseline 12minutes/day, 95% confidence interval (CI) -20.2 to 44.1]. In addition, there were no significant differences in total SF-36 or EORTC QLQ-LC13 scores from baseline to 4 weeks after surgery. Both groups had recovered their pre-operative walking distance 4 weeks after surgery, and there were no differences between the groups (mean difference in Incremental Shuttle Walk Test from baseline to 4 weeks after surgery (-26m, 95% CI -94.2 to 42.3). CONCLUSIONS: A hospital plus home exercise programme showed little benefit in unselected patients with NSCLC following surgery. Regardless of group allocation, the patients had recovered their pre-operative exercise tolerance levels by 4 weeks after surgery.

Evaluation of the in vitro activity of commercially available moxifloxacin and voriconazole eye-drops against clinical strains of Acanthamoeba.

PURPOSE: Acanthamoeba is an opportunistic pathogen which is the causal agent of a sight-threatening ulceration of the cornea known as "Acanthamoeba keratitis" (AK) and, more rarely, an infection of the central nervous system called "granulomatous amoebic encephalitis" (GAE). The symptoms of AK are non-specific, and so it can be misdiagnosed as a viral, bacterial, or fungal keratitis. Furthermore, current therapeutic measures against AK are arduous, and show limited efficacy against the cyst stage of Acanthamoeba. Moxifloxacin, a fourth generation fluoroquinolone, has been used with other drugs to treat GAE, but its efficacy as a treatment for AK is not known. Voriconazole has been used to treat AK; however, its cysticidal efficacy is not known. Both drugs are commercially available as eye-drops. The aim of this study was to evaluate the in-vitro activity of these eye-drops against Acanthamoeba compared to two reference drugs (chlorhexidine and amphotericin B) which are currently used to treat AK and GAE. METHODS: The sensitivity of two clinical and one type strain of Acanthamoeba to the commercial concentrations of the four drugs was evaluated with a colorimetric assay. Mature cysts were incubated with voriconazole to determine their sensitivity to this drug. The effects on cell proliferation and cell toxicity were determined using standard procedures with commercial kits. RESULTS: The four compounds were active against the Acanthamoeba strains in this study. Although it prevented encystation, moxifloxacin's amoebicidal activity was low. Voriconazole activity was greater than that of the other drugs, even at a concentration lower than in commercial eye drops. It was effective against cysts and decreased cell proliferation, with low cellular cytotoxicity. CONCLUSION: Voriconazole could be used against AK as a first-line treatment or in combination. Moxifloxacin is an interesting adjuvant to consider as it is effectively prevents encystation of the amoeba which often complicates infection resolution. In addition, moxifloxacin is effective in preventing secondary bacterial infections.

Oral transmission of Chagas disease: typing of Trypanosoma cruzi from five outbreaks occurred in Venezuela shows multiclonal and common infections in patients, vectors and reservoirs.

In Venezuela six episodes of oral transmission of Chagas disease (OChD) have been described, being the one reported in 2007 with a total of 103 people infected the largest worldwide. This work shows the use of three molecular markers (mini-exon gene and domains 24Sα and 18S of the ribosomal RNA) to characterize the infecting Trypanosoma cruzi strain of patients, reservoirs and vectors involved in five of the six OChD outbreaks. For this, 28 T. cruzi isolates were characterized by PCR, and the products of these reactions cloned and sequenced to reveal the existence of different TcI SL-IR genotypes. We also describe a new PCR assay able to discriminate between TcIb and TcId parasite populations. In summary, we have identified mostly parasites with the TcId haplotype and multiclonal populations with predominance of haplotype TcId (65.2%). Additionally, populations of haplotypes TcIb, TcIa and mixtures (TcId+TcIb, TcId+TcIa, TcIb+TcIa) are recurrent in samples obtained from children. The analysis of the SL-IR motif showed two clones depicting a different motif that could be an evidence for a possible hybrid haplotype between TcIa and TcIb (haplotype TcIa/Ib). Interestingly, in a single patient haplotype differences between T.cruzi isolates obtained pre and post-treatment were found. In conclusion, our findings show that in order to understand the pathogenic mechanisms involved in the orally acquired Chagas disease there is a need to join efforts to study T. cruzi haplotypes, their tissue tropisms and their susceptibility to chemoteraphy.

Muerte fetal intrauterina: ¿podemos actuar en su prevención? / Intrauterine fetal death: Can we act on prevention?

Objetivos: Análisis de los factores etiopatogénicos, maternos, fetales y placentarios, asociados a la muerte fetal intrauterina (MFIU). Análisis de los avances en el diagnóstico causal de la MFIU tras instauración de un protocolo específico. Métodos: Estudio descriptivo retrospectivo. Análisis de todos los casos de muerte fetal tardía en gestaciones únicas acontecidas en el Complejo Hospitalario Universitario de Vigo (2005- 2010). Resultados: Hubo 56 casos de muerte fetal tardía. De las gestantes estudiadas, 4 eran menores de 17 años y 19 mayores de 35 años, un 21,4 por ciento fumaban, el 60 por ciento tenía un IMC > 25 kg/m2 y el 18 por ciento no controlaron el embarazo. La patología materna predominante fue la tiroidea, mientras que las patologías gestacionales principales fueron diabetes gestacional, preeclampsia y amenaza de parto prematuro. El principal motivo de consulta fue la disminución de movimientos fetales. Se realizó estudio anatomopatológico placentario en el 82 por ciento y necropsia en el 73 por ciento. El porcentaje de causa desconocida en el grupo de no protocolo fue 20 por ciento y con protocolo 15 por ciento. Conclusión: El establecimiento de la causa de MFIU es difícil y en algunos casos no posible, aunque sí el reconocimiento de factores de riesgo. La implantación de un protocolo permitió un mayor acercamiento a la causa de la muerte fetal y mejor manejo posterior. La autopsia, el estudio de la placenta, los análisis citogenéticos y el estudio de trombosis materna son la base para el diagnóstico de MFIU.

Objective: Analysis of the pathogenetic maternal fetal and placental factors associated with stillbirth. Analysis of progress in the diagnoses of the cause of stillbirth after the establishment of a specific protocol. Methods: Retrospective descriptive study. Analysis of the cases of late fetal death in singleton pregnancies occurred at the University Hospital of Vigo during the period 2005-2010. Results: We found 56 late fetal deaths. Four were under 17 and 19 above 35 years old, 21percent were smokers, 60 percent had a BMI > 25 kg/m2 and 18 percent had not prenatal care. Thyroid disease was the most prevalent whereas gestational diabetes, preeclampsia and threatened preterm labour were the main obstetric pathologies. The main reason of consultation was decrease of fetal movements. Pathological exam of the placenta was made in 82 percent of cases and necropsy in 73 percent. Rate of unknown cause in the cases of no protocol was 20 percent while in the protocol group was 15 percent. Conclusion: The cause of intrauterine fetal death is difficult to establish. The recognition of certain risk factors is possible. The implementation of a study protocol allowed a better approach to the cause of fetal death and its management. Autopsy, placental examination, cytogenetic analysis, maternal thrombosis study was basic tests for the diagnosis of intrauterine fetal death.

Galectin and aldolase-like molecules are responsible for the specific IgE response in humans exposed to Dirofilaria immitis.

Dirofilaria immitis is the agent of the heartworm disease in canids and felids, and of pulmonary dirofilariosis in man. Like other filariae, D. immitis harbours endosymbion Wolbachia bacteriae. In this work we analyse the response of specific IgE antibodies against both D. immitis antigens and the Wolbachia surface protein (WSP) in two groups of persons living in an area of canine endemia, one presenting high levels of total IgE (group 1) and other with normal levels (group 2). Infections with D. immitis were demonstrated by the presence of specific IgG in 228 individuals(48.8%) of the group 1 and only in one of the group 2. Specific IgE antibody response against D. immitis antigens was detected only in individuals of the group 1. IgE response against WSP was not detected in any group. The IgE response was directed mainly against two molecules of 33 and 42 kDa of the antigenic extract of D. immitis. These molecules were identified by mass spectrometry as a galectin and an aldolase, respectively. Their possible role in the survival mechanisms of the parasite and their contribution to development of allergic reactions in individuals resident in areas with heartworm disease are discussed.

Identification of genotypes of Giardia intestinalis of human isolates in Egypt.

In Egypt, the genotyping study of Giardia intestinalis in human is limited. To determine the prevalence of G. intestinalis, faecal samples were collected from Egypt. Samples were concentrated using density gradient centrifugation. The samples were subjected to PCR and DNA sequence analysis for TPI gene. Prevalence of Giardia infection was 34.6% of 52 examined. DNA sequence showed that the Assemblage B was the most prevalent (80%) genotype, the 15% of the positive samples belonged to Assemblage E, and the 5% of them belonged to Assemblage A. Certainly, both genotypes A and B are highly common in human worldwide. However, up to now, Assemblage E had not been known to be infectious for humans. Therefore, this is the first time that Assemblage E is reported in human. However, further analyses of a second locus are required to confirm this result. The extent to which Giardia-infected cattle in Egypt might pose a risk of human infection is unknown.

The sera from individuals suffering from cutaneous leishmaniasis due to Leishmania brazilensis present antibodies against parasitic conserved proteins, but not their human counterparts.

Sera from individuals suffering from leishmaniasis have been shown to strongly react against conserved proteins from the parasite, such as ribosomal, histones and heat-shock proteins. Some of these proteins have also been described as immunogenic in several auto-immune syndromes, and the detection of antibodies against them is considered to be indicative of disorder of the immune system. In this paper, we investigate whether there is any relationship between the recognition of some conserved proteins from leishmania braziliensis by individuals suffering from cutaneous (CL) and mucocutaneous (MCL) leishmaniasis, and the recognition of the human homologues of these antigens found in sufferers from autoimmune diseases. Our findings reveal that the immune response generated during CL and MCL is elicited specifically by the parasitic histone H1 and Hsp70, since the CL and MCL sera do not react against their human counterparts. In addition, evidence is presented showing the specific recognition of human proteins by the autoimmune sera, showing only a weak cross-reaction with the most divergent regions of the parasitic proteins.

Raillietiella morenoi sp. n. (Pentastomida) from Gallotia atlantica (Peters and Doria, 1882) (Lacertidae) in the Canary Islands.

Raillietiella morenoi sp. n., a new cephalobaenid pentastomid found in the lungs of Gallotia atlantica collected in the Canary Islands, is described. The new species belongs to the sharp-tipped posterior-hook type. The annulus number, morphology, and dimensions of copulatory spicules and the dimensions of anterior and posterior hooks separate Raillietiella sp. n. from the other raillietiedid sharp-tipped posterior-hook species of small lizard parasites in Africa. The host character of endemic protected species of G. atlantica in Alegranza Island posed great difficulty in obtaining more parasite materials. However, more studies are required to state the variability of this species and its possible distribution in other species in the Canary Islands, as well as in other Gallotia spp.

Pathogenic Acanthamoeba strains from water sources in Jamaica, West Indies.

In 2004, samples of tap water and of river and sea water associated with human activities were collected in Jamaica, West Indies, and checked for free-living Acanthamoeba. The morphologies of the cysts and trophozoites observed and the results of PCR-based amplifications with a genus-specific primer pair were used to identify the Acanthamoeba isolates. The potential of each isolate as a human pathogen was then evaluated using thermotolerance and osmotolerance assays and two PCR-based assays for Acanthamoeba pathogenesis. Acanthamoeba were identified in 36.1%, 26.4% and 49.6% of the tap-, river- and sea-water samples collected, respectively. Pathogenic potential was shown by 60.0% of the Acanthamoeba strains isolated from tap water, 68.4% of the strains from river water, and 40.4% of the seawater strains. Sequencing of ribosomal DNA revealed the T1, T2, T4, T5, T7, T9 and T11 genotypes. Isolates of the T4 genotype were collected from tap, rain and sea water and, as expected, exhibited the most pathogenic traits; most were osmotolerant, thermotolerant and expressing extracellular serine protease. This is the first study of the occurrence and distribution of Acanthamoeba in water in the West Indies, and the results confirm the presence of potentially pathogenic strains in Jamaica.

Taenia spp.: 18S rDNA microsatellites for molecular systematic diagnosis.

The 18S rDNA gene of adult worms of Taenia parva found in Genetta genetta in the Iberian Peninsula and larval stages of T. pisiformis from the wild rabbit (Oryctolagus cuniculus) in Tenerife (Canary Islands) were amplified and sequenced. The sequences of the 18S rDNA gene of T. parva (1768 bp) and T. pisiformis (1760 bp) are reported for the first time (GenBank accession nos. AJ555167-AJ555168 and AJ555169-AJ555170, respectively). In 168 alignment positions microsatellites in the 18S rDNA of both taxa were detected for the first time (TGC in T. parva and TGCT in T. pisiformis) and differences in their sequences with different repetition numbers were observed. The use of nucleotide sequences of this gene in the resolution of systematic problems in cestodes is discussed with reference to the systematic status of Taenia spp. and mainly in human taeniids such as T. solium, T. saginata, and Asian human isolates of Taenia.

A specific primer pair for the diagnosis and identification of Acanthamoeba astronyxis by random amplified polymorphic DNA-polymerase chain reaction.

Random amplified polymorphic DNA (RAPD) is a useful tool for species identification. The obtained band patterns can be used for specific primer pair design that is useful for species identification. In this study, a distinctive 485-bp band in Acanthamoeba astronyxis band patterns was found, using the OPC20 primer (ACTTCGCCAC). The band specificity was confirmed by hybridization, using it as a probe, against all OPC20 amplifications from different Acanthamoeba species. Once the fragment was sequenced, we used it to design a specific primer pair that was useful for the identification of different isolates as A. astronyxis species.

Parasites of Columba livia (Aves: Columbiformes) in Tenerife (Canary Islands) and their role in the conservation biology of the laurel pigeons.

The prevalence and intensity of the parasites from 50 wild doves (Columba livia) from the city of Santa Cruz de Tenerife, in the island of Tenerife (Canary Archipelago), were studied. The following ectoparasites were found in apparently healthy pigeons (prevalences are shown in percentage (%) and mean intensities with their standard deviations): the acari Dermanyssus gallinae (De Geer, 1778) (6, 241.0 +/- 138.9) and Tinaminyssus melloi Fain, 1962 (10%, 218.3 +/- 117.3); the louses, Columbicola columbae Linnaeus, 1758 (100%, 111.4 +/- 76.8) and Campanulotes bidentatus Scopoli, 1763 (94%, 48.4 +/- 26.6); and the pigeon fly, Pseudolynchia canariensis Macquart, 1839 (36%, 6.2 +/- 1.6). The endoparasites we found were: a haemoprotozoan species, Haemoproteus columbae Kruze, 1890 (82%, 14.8 +/- 10.3 per 1000); coccidian oocysts, Eimeria sp. (50%, 0.2 x 10(3) +/- 1.7 x 10(3) per gr); a cestode species Raillietina micracantha (Fuhrmann, 1909) López Neyra, 1947 (44%, 12.3 +/- 9.4); and four nematode species, Tetrameres (Tetrameres) fissispina (Diesing, 1861) Travassos, 1915 (4%, 99.5 +/- 34.1), Synhimantus (Dispharynx) spiralis (Molin, 1858) (8%, 46.8 +/- 11.6), Ascaridia columbae (Gmelin, 1790) Travassos, 1913 (40%, 8.4 +/- 8.8) and Aonchotheca sp. (18%, 6.0 +/- 3.1). Several species detected in our study can be pathogens for C. bollii and C. junoniae, which are endemic pigeons of the Canary Islands, considered endangered species. Parasites (ectoparasites, protozoa and helminths) of C. livia found in Tenerife and others from wild and farm birds in the island were considered as healthy controls.

Molecular characterization of the Leishmania braziliensis L6 ribosomal protein.

By screening a Leishmania braziliensis complementary DNA library with a pool of sera from leishmaniasis patients, the gene coding for L6 ribosomal protein was isolated. The sequence, genomic organization, and transcription of this gene are described in this article. The sequence analysis of the L. braziliensis L6 gene shows a single open reading frame, which codes for a protein of 192 amino acids (aa) with a hypothetical molecular mass of 20.9 kDa. The protein exhibits significant sequence similarity to L6 ribosomal proteins from higher eukaryotes and yeast. Thus, the L. braziliensis L6 protein contains 4 functional motifs, which are located at equivalent positions in other L6 ribosomal proteins described previously. Interestingly, the L6 ribosomal protein from L. braziliensis contains a specific region of 14 aa and a tyrosine kinase motif, which is absent in human and C. elegans L6 protein. The locus coding the L. braziliensis L6 ribosomal protein is formed by 2 gene copies arranged in tandem and located in a chromosome of approximately 0.9. Mb. The genes are actively transcribed as 2 polyadenylated transcripts of approximately 1.15 and 0.85 kb, which differ in their steady-state level and stability.

Molecular systematics of several cyclophyllid families (Cestoda) based on the analysis of 18S ribosomal DNA gene sequences.

The sequences of the 18S small subunit ribosomal DNA of five species of cyclophyllidean cestodes from the families Davaineidae, Anoplocephalidae and Dilepididae were determined. A species of tetrabothridid was also sequenced. These 18S sequences were combined with other available eucestode sequences in GenBank. From the 1,838 sites in the alignment, 375 bp (20%) were excluded from the analysis due to alignment issues inferred by manual inspection. Phylogenetic trees were obtained by maximum parsimony, neighbour-joining distance and maximum likelihood methods. Analyses showed that Cyclophyllidea is monophyletic and separate from Tetrabothrius spp. Lyruterina nigropunctata, which is now included in the family Paruterinidae, is more closely related to davaineids of the genus Raillietina than Pseudidiogenes nana (Davaineidae). P. nana and Choanotaenia infundibulum (Dilepididae) derive from the Davaineidae (or Raillietininae). The two species of Taenia (T. parva and T. pisiformis) formed a monophyletic sister group to the Davaineidae and Anoplocephalidae. The systematics of the Paruterinidae have been problematic and our results suggest a review of this family including other species with paruterine organ located in other families. The position of the Idiogeninae as a subfamily in the Davaineidae is also reviewed.

Molecular and immunological characterization of L14 ribosomal protein from Leishmania braziliensis.

The isolation and molecular characterization of the gene coding for L14 ribosomal protein from L. braziliensis is described. There are 2 copies of the gene per haploid genome, repeated in a head-to-tail tandem orientation and located in a single chromosome of approximately 950 kb. Northern blot analyses indicate the presence of a single transcript of 0.95 kb which is up-regulated when parasites reach the stationary growth phase. L. braziliensis L14 gene codes for a 175 amino acid long polypeptide showing 75-83% sequence identity with L14 proteins from trypanosomatids and approximately 25% with its counterparts from higher eukaryotic organisms. L14 ribosomal proteins from trypanosomatids and higher eukaryotes share along their molecules a similar distribution pattern of theoretically functional domains. L. braziliensis L14 recombinant protein is not recognized by sera from cutaneous leishmaniasis patients. Immunization of mice with one dose of L14 recombinant protein and a second dose of L14 protein covalently linked to the HSP70 from Trypanosoma cruzi induces a high antibody level against this L14 protein, which is mostly of the IgG2a subtype, as well as a strong increase in splenocyte proliferation index.