Reduction in Plasmodium falciparum Pfk13 and pfg377 allele diversity through time in southern Vietnam.

BACKGROUND: Plasmodium falciparum has acquired resistance to artemisinin in Southeast Asia, with mutations in the P. falciparum Kelch-13 (Pfk13) gene associated with the resistance phenotype. The widespread use of Artemisinin-based combination therapy (ACT)s in Southeast Asia has led to the selection and spread of parasites carrying mutations in Pfk13. We characterised the allele diversity of Pfk13 and pfg377, an artemisinin-resistance neutral polymorphic gene, in parasite DNA extracted human blood from in southern Vietnam in 2003, 2012, 2015 and 2018. METHOD: This study was conducted in Bu Gia Map commune, Binh Phuoc province, Vietnam, from May 2018 to January 2019. Twenty-four samples from 2018 to 2019, 30 from 2003, 24 from 2012 and 32 from 2015 were analysed. Malaria-infected human blood was collected by finger-prick and used for molecular analysis. A nested-PCR targeting the small subunit ribosomal RNA gene was used for Plasmodium species identification, followed by amplification and nucleotide sequencing of Pfk13 and region 3 of pfg377. Archived blood samples collected in the same region in 2012 and 2015 were also analysed as above for comparison. RESULTS: The genetic diversity of Pfk13 and pfg377 was lower in 2018-2019 compared to 2012 and 2015. The number of distinct Pfk13 mutants decreased from three in 2012 and 2015, P553L, V568G and C580Y, to one, C580Y in 2018-2019. In 2018-2019, the frequency of C580Y mutant strains was 71% (17/24 isolates). All samples were wild type in 2003. In 2012 and 2015, there were single-strain infections as well as co-infections with two mutant strains or with mutant and wild strains, whereas there were no co-infections in 2018. pfg377 allele diversity decreased from five alleles in 2012 to two alleles in 2018-2019. CONCLUSION: The genetic diversity of P. falciparum was reduced at the two genetic loci surveyed in this study, Pfk13 and pfg377. In the case of the former gene, we observed an increase in the prevalence of parasites carrying the C580Y gene, known to confer reduced susceptibility to ACTs. The reduction in the diversity of pfg377 may be linked to the clonal expansion of parasite strains carrying the C580Y mutation, leading to an overall reduction in parasite genetic diversity across the population.

Prevalence of human and non-human primate Plasmodium parasites in anopheline mosquitoes: a cross-sectional epidemiological study in Southern Vietnam.

BACKGROUND: Human malaria is a major threat in rural communities of central Vietnam. Anopheles dirus and Anopheles minimus species are critical malaria vectors in Vietnam, which transmit Plasmodium parasites. However, the entomological aspects of malaria transmission in some of the central provinces of Vietnam remain unexplored. Hence, a cross-sectional entomological survey was carried out to identify the malaria vector species and the transmission of Plasmodium parasites in seven endemic provinces of Vietnam. METHODS: Mosquitoes were collected from seven provinces, Gia Lai, Khanh Hoa, Phu Yen, Ninh Thuan, Binh Thuan, Dong Nai, and Binh Phuoc. The collection was conducted for four to eight consecutive nights using three established methods, indoor and outdoor human landing catches and light trap method. Nested-PCR analysis was performed to detect the Plasmodium species in the separated thorax and the abdomen of the individual mosquitoes. RESULTS: A total of 2278 mosquitoes belonging to one of the four species of anopheline mosquitoes, An. dirus, An. maculatus, An. aconitus, and An. minimus were collected. Among the collected mosquitoes, 1398 were analysed using nested-PCR, of which, 40 mosquitoes were positive for Plasmodium parasites. Most of these parasites were detected in the samples from the thorax region, followed by the abdominal portion. The parasites were detected in both the thorax and abdomen of An. dirus. Seven species of Plasmodium parasites were detected during the analysis, of which, Plasmodium inui was the most common species, followed by Plasmodium falciparum, Plasmodium vivax, Plasmodium cynomolgi, Plasmodium coatneyi, Plasmodium knowlesi, and Plasmodium fieldi. Out of the 49 positive samples, 12 showed mixed infections. Co-infection of P. inui with human and other non-human primate Plasmodium species was common. CONCLUSIONS: This study demonstrated the presence of human and non-human primate Plasmodium infection in An. dirus, a predominant malarial vector. Further, we showed that An. maculatus and An. minimus species also take part in malarial transmission. This might potentially lead to an alarming situation conducive for the emergence of novel zoonotic malaria.

Human infection with Plasmodium knowlesi on the Laos-Vietnam border.

BACKGROUND: Border malaria in the Greater Mekong region of Southeast Asia poses a serious threat to the health of the ethnic minority populations of the region. Traditionally thought to be caused primarily by the malaria parasites Plasmodium falciparum and Plasmodium vivax, recently a zoonotic parasite, Plasmodium knowlesi, has been identified in some countries of the region. The presence of this parasite poses a challenge to malaria control programmes, as it is maintained in a zoonotic reservoir of forest-dwelling macaque monkeys. METHODS: A cross-sectional malaria parasite species prevalence survey was conducted along the Laos-Vietnam border in the central part of the two countries. Human blood samples were collected from Savannakhet in Laos and Quang Tri in Vietnam between August and October 2010 and assayed for the presence of human malaria parasite species and P. knowlesi. A PCR targeting the 18S small subunit ribosomal RNA gene and circumsporozoite protein gene was used for Plasmodium species identification. RESULTS: Nine cases of P. knowlesi were detected by PCR in blood samples from the Laos side and three from the Vietnam side. All P. knowlesi infections were found in co-infection with P. vivax, with some triple infections of P. knowlesi, P. vivax and P. falciparum detected in Laos. Phylogenetic analysis of these parasites suggests that P. knowlesi is circulating in the Laos-Vietnam border region. CONCLUSION: This report shows that P. knowlesi is transmited on both sides of the Vietnam-Laos border. Continued monitoring of the range and prevalence of P. knowlesi on both the sides of Laos-Vietnam border is of importance to the National Malaria Control Programmes of both countries.

Molecular epidemiology of mosquitoes for the transmission of forest malaria in south-central Vietnam.

Human infection caused by non-human primate malarial parasites, such as Plasmodium knowlesi and Plasmodium cynomolgi, occurs naturally in Southeast Asian countries, including Vietnam. Members of the Anopheles dirus species complex are known to be important vectors of human malarial parasites in the forested areas of southern and central Vietnam, including those in Khanh Phu commune and Khanh Hoa Province. Recent molecular epidemiological studies in Vietnam have reported cases of co-infection with Plasmodium falciparum, Plasmodium vivax, Plasmodium malariae, and P. knowlesi in An. dirus. The commonly found macaques in the forest in the forested areas are suspected to be bitten by the same An. dirus population that bites humans. A recent epidemiological study identified six species of malarial parasites in sporozoite-infected An. dirus using polymerase chain reaction, of which P. vivax was the most common, followed by P. knowlesi, Plasmodium inui, P. cynomolgi, Plasmodium coatneyi, and P. falciparum. Based on a gametocyte analysis, the same allelic gametocyte types were observed in both humans and mosquitoes at similar frequencies. These observations suggest that people who stay overnight in the forests are frequently infected with both human and non-human primate malarial parasites, leading to the emergence of novel zoonotic malaria. Moreover, it is suggested that mosquito vector populations should be controlled and monitored closely.

Detection of the Plasmodium falciparum Kelch-13 gene P553L mutation in sporozoites isolated from mosquito salivary glands in South-Central Vietnam.

BACKGROUND: Plasmodium falciparum has developed resistance against artemisinin in Southeast Asia. Mutations in the P. falciparum Kelch-13 (Pfk13) gene are associated with artemisinin resistance in vitro and in vivo. We investigated the prevalence of mutations in PfK13 from sporozoite-stage parasites isolated from the salivary glands of Anopheles dirus mosquitoes. METHODS: Mosquitoes were caught by human-landing catches at two locations within the Khanh Phu commune, South-Central Vietnam. Identification of Anopheles species was performed based on morphological features and nucleotide sequence analysis. Sporozoite-infected salivary glands were stored on filter paper and at 4-6 °C. A nested-PCR targeting the small subunit ribosomal RNA gene was used for Plasmodium species identification. Pfk13 was amplified by nested PCR, and subjected to nucleotide sequencing. RESULTS: Five of 33 P. falciparum sporozoite samples carried the P553L mutation at the PfK13 locus. This mutation has been recorded previously in Vietnam, but not in Khanh Hoa province, were surveys of K13 polymorphism have not previously been carried out. CONCLUSION: These results demonstrate the utility of mosquito-stage malaria parasite samples for studies on the molecular epidemiology of drug resistance.

Molecular epidemiology of rotavirus gastroenteritis in Central Kenya before vaccine introduction, 2009-2014.

Between July 2009 and June 2014, a total of 1,546 fecal specimens were collected from children <5 years of age with acute gastroenteritis admitted to Kiambu County Hospital, Central Kenya. The specimens were screened for group A rotavirus (RVA) using ELISA, and RVA-positive specimens were subjected to semi-nested RT-PCR to determine the G and P genotypes. RVA was detected in 429/1,546 (27.5%) fecal specimens. RVA infections occurred in all age groups <59 months, with an early peak at 6-17 months. The infections persisted year-round with distinct seasonal peaks depending on the year. G1P[8] (28%) was the most predominant genotype, followed by G9P[8] (12%), G8P[4] (7%), G1P[4] (5%), G9P[4] (4%), and G12P[6] (3%). In the yearly change of G and P genotypes, a major shift from G9P[8] to G1P[8] was found in 2012. Phylogenetic analysis of the nucleotide sequences of the VP7 and VP4 genes of seven strains with unusual G8 or P[6] showed that the VP7 nucleotide sequences of G8 were clustered in lineage 6 in which African strains are included, and that there are at least two distinct VP4 nucleotide sequences of P[6] strains. These results represent basic data on RVA strains circulating in this region before vaccine introduction. J. Med. Virol. 89:809-817, 2017. © 2016 Wiley Periodicals, Inc.

Humans frequently exposed to a range of non-human primate malaria parasite species through the bites of Anopheles dirus mosquitoes in South-central Vietnam.

BACKGROUND: Recent studies have described natural human infections of the non-human primate parasites Plasmodium knowlesi and Plasmodium cynomolgi. In Southeast Asia, mosquitoes of the Anopheles leucosphyrus group bite both humans and monkeys in the forest and thus offer a possible route for Plasmodium species to bridge the species barrier. In this study we analysed the species composition of malarial sporozoites infecting the salivary glands of Anopheles dirus in order to determine their potential role as bridge vectors of Plasmodium parasites from monkeys to humans. METHODS: Mosquitoes were collected in the forest and forest fringe area of Khanh Phu commune by human-baited landing collection. Anopheles species were determined on the basis of morphologic features. Sporozoite-infected salivary glands were applied to filter paper and dried in an ambient atmosphere, before storage in closed vials at 4-6 °C. Detection and identification of Plasmodium species in salivary glands were carried out by nested-PCR of the small subunit ribosomal RNA gene. RESULTS: Six species of Plasmodium parasites were detected by PCR, of which P. vivax was the most common, followed by P. knowlesi, P. inui, P. cynomolgi, P. coatneyi and P. falciparum. Twenty-six of the 79 sporozoite infected mosquitoes showed multiple infections, most of which were a combination of P. vivax with one or more of the non-human primate Plasmodium species. CONCLUSIONS: These results suggest that humans overnighting in this forest are frequently inoculated with both human and non-human primate malaria parasites, leading to a situation conducive for the emergence of novel zoonotic malaria.

Detection of Plasmodium knowlesi DNA in the urine and faeces of a Japanese macaque (Macaca fuscata) over the course of an experimentally induced infection.

BACKGROUND: Diagnostic techniques based on PCR for the detection of Plasmodium DNA can be highly sensitive and specific. The vast majority of these techniques rely, however, on the invasive sampling of blood from infected hosts. There is, currently, considerable interest in the possibility of using body fluids other than blood as sources of parasite DNA for PCR diagnosis. METHODS: Urine and faeces were obtained from a Plasmodium knowlesi infected-Japanese macaque (Macaca fuscata) over the course of an experimentally induced infection. P. knowlesi DNA (PkDNA) extracted from urine and faeces were monitored by nested PCR targeting the P. knowlesi specific cytochrome b (cytb) gene. RESULTS: Urinary PkDNA was detected on day 2, but was not amplified using DNA templates extracted from the samples on day 4, day 5 and day 6. Subsequently, urinary PkDNA was detected from day 7 until day 11, and from day 20 until day 30. PkDNA in faeces was detected from day 7 until day 11, and from day 20 until day 37. Moreover, real-time quantitative PCR showed a remarkable increase in the amount of urinary PkDNA following anti-malarial treatment. This might have been due to the release of a large amount of PkDNA from the degraded parasites as a result of the anti-malarial treatment, leading to excretion of PkDNA in the urine. CONCLUSIONS: The cytb-PCR system using urine and faecal samples is of potential use in molecular epidemiological surveys of malaria. In particular, monkey faecal samples could be useful for the detection of zoonotic primate malaria in its natural hosts.

Whole genomic analysis of human G12P[6] and G12P[8] rotavirus strains that have emerged in Kenya: identification of porcine-like NSP4 genes.

G12 rotaviruses are globally emerging rotavirus strains causing severe childhood diarrhea. However, the whole genomes of only a few G12 strains have been fully sequenced and analyzed, of which only one G12P[4] and one G12P[6] are from Africa. In this study, we sequenced and characterized the complete genomes of three G12 strains (RVA/Human-tc/KEN/KDH633/2010/G12P[6], RVA/Human-tc/KEN/KDH651/2010/G12P[8], and RVA/Human-tc/KEN/KDH684/2010/G12P[6]) identified in three stool specimens from children with acute diarrhea in Kenya, Africa. On whole genomic analysis, all three Kenyan G12 strains were found to have a Wa-like genetic backbone: G12-P[6]-I1-R1-C1-M1-A1-N1-T1-E1-H1 (strains KDH633 and KDH684) and G12-P[8]-I1-R1-C1-M1-A1-N1-T1-E1-H1 (strain KDH651). Phylogenetic analysis showed that most genes of the three strains examined in this study were genetically related to globally circulating human G1, G9, and G12 strains. Of note is that the NSP4 genes of strains KDH633 and KDH684 appeared to be of porcine origin, suggesting the occurrence of reassortment between human and porcine strains. Furthermore, strains KDH633 and KDH684 were very closely related to each other in all the 11 gene segments, indicating derivation of the two strains from a common origin. On the other hand, strain KDH651 consistently formed distinct clusters of 10 of the 11 gene segments (VP1-2, VP4, VP6-7, and NSP1-5), indicating a distinct origin of strain KDH651 from that of strains KDH633 and KDH684. To our knowledge, this is the first report on whole genome-based characterization of G12 strains that have emerged in Kenya. Our observations will provide important insights into the evolutionary dynamics of emerging G12 rotaviruses in Africa.

DNA from pre-erythrocytic stage malaria parasites is detectable by PCR in the faeces and blood of hosts.

Following the bite of an infective mosquito, malaria parasites first invade the liver where they develop and replicate for a number of days before being released into the bloodstream where they invade red blood cells and cause disease. The biology of the liver stages of malaria parasites is relatively poorly understood due to the inaccessibility of the parasites to sampling during this phase of their life cycle. Here we report the detection in blood and faecal samples of malaria parasite DNA throughout their development in the livers of mice and before the parasites begin their growth in the blood circulation. It is shown that parasite DNA derived from pre-erythrocytic stage parasites reaches the faeces via the bile. We then show that different primate malaria species can be detected by PCR in blood and faecal samples from naturally infected captive macaque monkeys. These results demonstrate that pre-erythrocytic parasites can be detected and quantified in experimentally infected animals. Furthermore, these results have important implications for both molecular epidemiology and phylogenetics of malaria parasites. In the former case, individuals who are malaria parasite negative by microscopy, but PCR positive for parasite DNA in their blood, are considered to be "sub-microscopic" blood stage parasite carriers. We now propose that PCR positivity is not necessarily an indicator of the presence of blood stage parasites, as the DNA could derive from pre-erythrocytic parasites. Similarly, in the case of molecular phylogenetics based on DNA sequences alone, we argue that DNA amplified from blood or faeces does not necessarily come from a parasite species that infects the red blood cells of that particular host.

Whole genomic analysis of a porcine-like human G5P[6] rotavirus strain isolated from a child with diarrhoea and encephalopathy in Japan.

An unusual rotavirus strain, Ryukyu-1120, with G5P[6] genotypes (RVA/Human-wt/JPN/Ryukyu-1120/2011/G5P[6]) was identified in a stool specimen from a hospitalized child aged 4 years who showed diarrhoea and encephalopathy. In this study, we sequenced and characterized the complete genome of strain Ryukyu-1120. On whole genomic analysis, this strain was found to have a unique genotype constellation: G5-P[6]-I5-R1-C1-M1-A8-N1-T1-E1-H1. The VP6 and NSP1 genotypes I5 and A8 are those commonly found in porcine strains. Furthermore, phylogenetic analysis indicated that each of the 11 genes of strain Ryukyu-1120 appeared to be of porcine origin. Thus, strain Ryukyu-1120 was found to have a porcine rotavirus genetic backbone and is likely to be of porcine origin. To our knowledge, this is the first report of whole-genome-based characterization of the emerging G5P[6] strains in Asian countries. Our observations will provide important insights into the origin of G5P[6] strains and the dynamic interactions between human and porcine rotavirus strains.

Sudden death from systemic rotavirus infection and detection of nonstructural rotavirus proteins.

A 2.5-year-old girl died suddenly during the course of rotavirus gastroenteritis. The autopsy showed encephalopathy with rotavirus systemic infection. Here, we provide evidence of rotavirus replication in multiple organs. Our findings clarify that rotavirus infection in children can extend beyond the intestinal tract through viremia.

Modification of the trypsin cleavage site of rotavirus VP4 to a furin-sensitive form does not enhance replication efficiency.

The infectivity of rotavirus (RV) is dependent on an activation process triggered by the proteolytic cleavage of its spike protein VP4. This activation cleavage is performed by exogenous trypsin in the lumen of the intestines in vivo. Here, we report the generation and characterization of a recombinant RV expressing cDNA-derived VP4 with a modified cleavage site (arginine at position 247) recognized by endogenous furin as well as exogenous trypsin. Unexpectedly, the mutant virus (KU//rVP4-R247Furin) was incapable of plaque formation without an exogenous protease, although the mutant VP4s on virions were efficiently cleaved by endogenous furin. Furthermore, KU//rVP4-R247Furin showed impaired infectivity in MA104 and CV-1 cells even in the presence of trypsin compared with the parental virus carrying authentic VP4 (KU//rVP4). Although the total titre of KU//rVP4-R247Furin was comparable to that of KU//rVP4, the extracellular titre of KU//rVP4-R247Furin was markedly lower than its cell-associated titre in comparison with that of KU//rVP4. In contrast, the two viruses showed similar growth in a furin-defective LoVo cell line. These results suggest that intracellular cleavage of VP4 by furin may be disadvantageous for RV infectivity, possibly due to an inefficient virus release process.

Co-infections of Plasmodium knowlesi, P. falciparum, and P. vivax among Humans and Anopheles dirus Mosquitoes, Southern Vietnam.

A single Anopheles dirus mosquito carrying sporozoites of Plasmodium knowlesi, P. falciparum, and P. vivax was recently discovered in Khanh Phu, southern Vietnam. Further sampling of humans and mosquitoes in this area during 2009-2010 showed P. knowlesi infections in 32 (26%) persons with malaria (n = 125) and in 31 (43%) sporozoite-positive An. dirus mosquitoes (n = 73). Co-infections of P. knowlesi and P. vivax were predominant in mosquitoes and humans, while single P. knowlesi infections were found only in mosquitoes. P. knowlesi-co-infected patients were largely asymptomatic and were concentrated among ethnic minority families who commonly spend nights in the forest. P. knowlesi carriers were significantly younger than those infected with other malaria parasite species. These results imply that even if human malaria could be eliminated, forests that harbor An. dirus mosquitoes and macaque monkeys will remain a reservoir for the zoonotic transmission of P. knowlesi.

In vivo and in vitro gametocyte production of Plasmodium falciparum isolates from Northern Thailand.

Understanding why some malaria-infected individuals are infective to mosquitoes while others are not, is of great importance when considering interventions to stop malaria transmission. Whether gametocytes are produced in every individual infected with Plasmodium falciparum remains unclear. Using a highly sensitive reverse transcription (RT)-PCR assay, we attempted to detect gametocyte-specific mRNA transcripts in isolates from Thai patients which newly adapted to continuous in vitro culture. We then compared the allelic types of the pfg377 gene between patient blood and culture-adapted parasites in order to determine whether the same parasite lines were producing gametocytes in vivo and in vitro. Transcripts of pfg377 were detected in all parasite isolates and in the corresponding cultured isolates, revealing that all patients had gametocytes circulating in their blood at the time of sampling. For isolates in continuous in vitro culture, there was a match between pfg377 allelic types detected by PCR from genomic DNA (and thus indicative of the dominant allelic type of asexual parasites) and those detected by RT-PCR of mRNA (gametocyte-specific), whereas in freshly isolated patient blood there were some differences between the asexual parasite allelic type and that of the gametocytes in the same infection. Seven isolates contained asexual stage parasites harbouring pfg377 alleles that were not detectable in gametocytes from the same infections, suggesting that some clones were not producing gametocytes at the time of sampling, or that they were below the level of detection.

Anopheles dirus co-infection with human and monkey malaria parasites in Vietnam.

The feasibility of identifying parasite DNA and specific mRNAs from wild-caught Anopheles dirus mosquitoes was assessed using dried mosquito salivary glands preserved on filter paper. We were able to detect Plasmodium falciparum, Plasmodium vivax, Plasmodium malariae and Plasmodium knowlesi DNA by conventional PCR and, furthermore, detected P. falciparum gametocyte-specific genes, pfg377 and pfs16 mRNA, P. knowlesi circumsporozoite protein (CSP) and sporozoite surface protein 2 (SSP2) mRNA by reverse transcription-PCR. Using this technique, we were able to confirm the presence of P. vivax, P. falciparum and P. knowlesi in one particular wild-caught mosquito. These results indicate that P. knowlesi may be transmitted by the primary human malaria vector in forested areas in Vietnam. This study also shows that the preservation of mosquito salivary glands on filter paper, and the down-stream extraction of parasite DNA and RNA from those, offers a powerful resource for molecular epidemiological studies on malaria.

Effect of osteopontin on diarrhea duration and innate immunity in suckling mice infected with a murine rotavirus.

We studied the role of osteopontin (OPN) in host responses against rotavirus (RV) infection. OPN knockout (OPN-KO) suckling mice were more susceptible to RV (strain EW) infection and showed prolonged diarrhea duration compared to wild-type (WT) suckling mice. OPN in the small intestine of WT mice was expressed after 48 h post-infection. On day 2 postinfection, mRNA levels of interleukin-1beta, tumor necrosis factor-alpha, and interleukin-15 in OPN-KO mice were lower than in WT mice, although mRNA expression of Th-1- and Th-2-related cytokines in the small intestine were nearly the same between OPN-KO and WT mice. These results suggested that OPN is involved in innate responses against RV infection.

A dried blood sample on filter paper is suitable for detecting Plasmodium falciparum gametocytes by reverse transcription polymerase chain reaction.

The detection of gametocytes in human peripheral blood is one of the most important measures in a malaria survey. We attempted to detect gametocytes of Plasmodium falciparum by reverse transcription polymerase chain reaction (RT-PCR) of dried blood on filter paper. On field samples analysis, the specific RT-PCR products for region 3 of pfg377 mRNA were observed in 67 of 131 falciparum malaria patients. The minimum detection level of RT-PCR-positive samples was 0.03 gametocytes/microl on quantitative real-time RT-PCR. Gametocyte positive rate was not dependent on sex or age. A higher frequency of gametocytes was found in single P. falciparum infection than in mixed species infection (P<0.01). In this study, 47 of the 131 patients were asymptomatic. Eighteen of these 47 patients showed pfg377 mRNA expression. Moreover, four alleles of region 3 of pfg377 were detected in pfg377 mRNA-positive patients and 13 of 67 pfg377 mRNA-positive patients carried more than one gametocyte-producing clone. These results suggest that dried blood on filter paper is a useful for a molecular epidemiologic study of malaria transmission and gametocyte-targeted control.

Osteopontin is involved in Th1-mediated immunity against Plasmodium falciparum infection in a holoendemic malaria region in Vietnam.

We examined the role of osteopontin (OPN) in immunity against Plasmodium falciparum infection. We measured the mRNA levels for OPN and several cytokines in RNA preparations extracted from dried blood on filter paper obtained from falciparum malaria patients in Vietnam. Expression of OPN mRNA was detected in 134 of 161 patients. The expression of both interleukin-12 p40 and interferon-gamma mRNAs in the group positive for OPN mRNA was significantly higher than that in the group negative for OPN mRNA. The level of parasitemia in the OPN mRNA-positive group was much lower than that in the negative one. These results suggest that OPN might suppress multiplication of the parasites through T helper 1 cells-mediated immune responses.

Osteopontin participates in Th1-mediated host resistance against nonlethal malaria parasite Plasmodium chabaudi chabaudi infection in mice.

Osteopontin (OPN) knockout mice (OPN-KO mice) died of Plasmodium chabaudi chabaudi infection, although wild-type (WT) mice had self-limiting infections. OPN was detected in the WT mice at 2 days postinfection. OPN-KO mice produced significantly smaller amounts of interleukin-12 and gamma interferon than WT mice produced. These results suggested that OPN is involved in Th1-mediated immunity against malaria infection.